Comparison of the effect of sodium bicarbonate and glycine air polishing systems on tooth surface roughness: An atomic force microscopic analysis

Abstract

BACKGROUND:

A variety of prophylactic materials are used in the dental office for the removal of stains and calculus.

OBJECTIVE:

To evaluate tooth surface changes caused by the application of air abrasive powders (sodium bicarbonate, SBAP and glycine air powder, GPAP) along with scaling and root planing (SRP), under atomic force microscope (AFM) and to analyze the histological soft tissue changes caused by these agents, using light microscopy.

METHODS:

This study was conducted in two phases: in vitro and in vivo. In the in vitro phase, hard tissue analysis was done under AFM following air powder polishing. Eighteen extracted teeth were chosen. SRP and tooth sectioning were carried out. Subsequently, each section of the tooth was mounted on a glass plate with self-cure acrylic resin and air polished using SBAP and GPAP. In the vivo phase, the soft tissue was analyzed under a light microscope for surface roughness. A biopsy specimen was taken from patients who had received phase I therapy, and flap surgery was planned using a modified Widman flap technique.

RESULTS:

This study compared surface changes in enamel and cementum, under AFM, as indicated by R ${}_{A}$ after SRP, SRP and SBAP, and SRP and GPAP; comparisons were then drawn across the three groups. The mean AFM values were 108.5 and 144.7, 102.7 and 81.7, and 95.6 and 7.4, at the crown and root, for SRP, SRP and SBAP, and SRP and GPAP interventions, respectively. GPAP was the least rough on soft tissues.

CONCLUSION:

SBAP and GPAP were better than hand instrumentation as indicated by AFM and histological section analysis.

Keywords

Periodontics dentistry tooth surface atomic force microscope (AFM)

1. Introduction

Periodontitis is a chronic inflammatory disease caused by a plethora of etiological factors, including approximately 400 oral bacterial species that cause pathogenesis and pocket formation; this leads to loosening and ultimately to tooth loss. Various anatomical factors play an essential role for the accumulation of bacteria in both supragingival and subgingival areas. Surface roughness [1] acts as a nidus for the growth and development of bacterial colonies; thus, it requires special consideration. Various mechanical aids, such as hand instrumentation, ultrasonic scalers, and polishing systems (rubber cups and brush cones) with abrasive pastes, are used in periodontal therapy; these have their own advantages and disadvantages. At the same time, the frequent use of mechanical debridement is time consuming and may produce microabrasions over the tooth surface[2]. As an alternative, an air polishing system, with air abrasive powder, water, and pressurized air could aide routine professional cleaning procedures [3].

When considering air polishing (AP) techniques, the abrasive particle size, angulation, time, distance, and its effects on the tooth and soft tissue surfaces need to be evaluated. To date, many polishing powders have been used, of which sodium bicarbonate and glycine air powders are the most common. Polishing agents should cause minimal surface changes on the tooth, including roughness or smoothness, which are directly related with bacterial adhesion. Thus, it is essential to evaluate the structural integrity of the hard tissues after polishing. This allows for the measurement of the absolute loss of material from the tooth surface and the amount of roughness generated by the procedure. In the literature, there are many physical techniques that measure the surface roughness, including laser scanners, profilometers, scanning electron microscopes, and atomic force microscopes (AFM). Various studies [4, 5, 6] have been conducted using these techniques, among which the analysis of tooth surfaces with AFM are the least explored. One recognized advantage of these techniques is their ability to allow the characterization of large areas containing both untreated and AP treated regions. This makes it possible to measure the resulting defect depth and the absolute loss of material, and thus evaluate the integrity of dental structures [7]. However, laser scanners and profilometers do not permit high-resolution measurement of surface roughness. On the contrary, AFM [8, 9] can provide three dimensional images of the tooth surface with high resolution on a nanometer scale at a particular point without any manipulation of the sample. Also, since AFM is not destructive, the same samples can be sequentially imaged when subjected to various treatments, or they can be further integrated with other destructive surface analytical techniques for correlational studies. Therefore, AFM gives us a window into the nanoscale level of tooth structure. AFM is based upon the cantilever/tip assembly, which interacts with the sample. This assembly is also commonly referred to as the probe. The up/down and side-to-side motion of the AFM tip, as it scans along the surface, is monitored through a laser beam reflected off the cantilever. Various studies [4, 5, 8, 9] have been conducted to study the surface roughness of a tooth using various variables following AFM but to the best of our knowledge no study has evaluated the changes on the tooth surface following AP using AFM and correlated it with soft tissue changes using histological sections. Thus, our study aimed to evaluate the tooth surface changes caused by applying air abrasive powders (sodium bicarbonate and glycine air powder) along with scaling and root planing, under AFM and to analyze histological soft tissue changes caused by these agents, using light microscopy.

2. Methods

The present study was conducted at the Department of Periodontics, K.S.R. Institute of Dental Science and Research, Tiruchengode, India and was approved by the institute’s ethics committee (ethical clearance number 008/KSRIDSR/EC/2011). This study was conducted in two phases: in vitro and in vivo. In the in vitro phase, hard tissue analysis was done following AP using AFM. In the in vivo phase, soft tissue analysis was done using histological sections under a light microscope. Each phase was planned and carried out meticulously.

3. In vitro phase – Hard tissue analysis

3.1 Tooth selection and preparation

In this phase, the first step was tooth selection and its preparation for analysis. This step was conducted in line with previous studies [10] and was performed using nanohybrid composite resin that was air polished with either sodium bicarbonate powder or glycine powder. Initially a total of 18 extracted, non-carious premolars (for orthodontic purposes) were selected and stored in physiological saline and used within a week. All teeth were chosen after being extracted and examined for gross surface irregularities as they can interfere with AFM analysis. On each selected tooth, markings were done to demarcate the crown and root. Following this, SRP and sectioning were carried out. Subsequently, each tooth section was mounted on a glass plate with self-cure acrylic resin and air polished. Care was taken while sectioning and mounting; the topography of the surface to be examined was then visualized (Fig. 1a–c).

Figure 1.

(a) Selected teeth with marking for demarcation of the crown and root portion; (b), supragingival and subgingival scaling and root planing; (c), air polishing of the sectioned teeth

3.2 Air-polishing process

Air polishing was performed using a standard air-polishing unit (Air-Flow, EMS), installed according to the manufacturer’s instructions. The instrument nozzle was kept at 45 degree angle to the slide surface. Spraying distance was kept constant at 4 mm by holding the nozzle with the analyzing surface, and the application time was 5 seconds for each surface.

There were 3 groups with 6 teeth per group: (i) Group I (Control), Supragingival scaling and root planing (SRP); (ii) Group II, Supragingival scaling and sodium bicarbonate air polishing (SRP and SBAP); (iii) Group III, Supragingival scaling and glycine air polishing (SRP and GPAP).

There were 6 teeth in each group, 2 sites on the crown (designated as crown 1 and crown 2) and two sites on the root (designated as root 1 and root 2) (cementum) were randomly identified and AP was done with sodium bicarbonate in group II and Glycine in group III. The instrument’s powder chamber was refilled after each air-polishing period to ensure maximum reproducibility of powder emission. To standardize the process, abrasive particle size, angulation, time, and distance were kept the same for all samples.

3.3 AFM measurements

In vitro assessment of tooth surface changes by AFM was done in the Department of Nanoscience and Technology, K.S.R. College of Science and Technology, Tiruchengode. The relative height maps of the sample surfaces, both for controls and AP treated specimens, were acquired with the help of probes. The probes used were in contact mode with the Nanosurf 2 EC scan at a constant force mode of 16 nano newton. The image magnification used was 5 nanometers with the Nanosurf software and was also recorded. The surface roughness of each specimen was evaluated as the root mean square (RMS) value R ${}_{A}$ of the distribution of heights in the three-dimensional AFM topographic images

Figure 2.

(a) Images and graphs representing root mean square area (R ${}_{a}$ ) on nano scale for supragingival and subgingival scaling and root planing. (b) Images and graphs representing root mean square area (R ${}_{a}$ ) on nano scale for supragingival and subgingival scaling and root planing with sodium bicarbonate air polishing. (c) Images and graphs representing root mean square area (R ${}_{a}$ ) on nano scale for supragingival and subgingival scaling and root planing with glycine air polishing.

3.4 Soft tissue analysis

For the in vivo aspect, a total of 6 patients were selected, and informed consent was taken from all patients. Ethical clearance was also obtained from the institutes’ ethical committee. Patients were recruited from the Department of Periodontology, K.S.R. Institute of Dental Science and Research, Tiruchengode. The recruitment period was from January 2013 to March 2013. Patients who received phase I therapy and presented 4–6 weeks later with 5 mm or greater probing depth on at least four teeth which were selected for the Modified Widman flap procedure, were included in the study. Patients under 18 years and pregnant women were excluded. In the 6 selected patients after supragingival scaling and root planing, 12 randomly selected sites per patient were divided into three groups (4 sites/group), on the basis of planned interventions with air polishing agents under observation. Following this, gingival tissue biopsy was carried out to study the soft tissue under a microscope.

Group I received supragingival scaling and root planing (SRP); Group II received supragingival scaling and sodium bicarbonate air polishing (SRP and SBAP); Group III received Supragingival scaling and glycine air polishing (SRP and GPAP).

3.4.1 Group I

For each patient supragingival scaling and root planing was done using a sharp Gracey curette, hard tissue debridement was performed until no plaque was visible on the instrument’s working end when it was retrieved from the periodontal pocket. Following this, internal bevel incision was carried out on the gingival soft tissue biopsy which included the sulcular and junctional epithelium from the selected sites (size 2 mm). Subsequently, the tissue was stored in formalin solution to avoid tissue shrinkage

3.4.2 Group II

Similar to group I, after the supragingival scaling and root planing, in group II on the selected sites sodium bicarbonate air polishing was carried out. Standardization protocols were strictly followed with the instrumentation. A distance of 4 mm with a 5 second duration for each tooth surface was maintained, and the nozzle of the instrument was angled at 45 degree to the surface. Following this, a biopsy was carried out and the tissue was stored in formalin solution. Moreover, to avoid the exposure of the gingival tissues adjacent to the tooth being investigated, the surrounding soft tissues were covered with tinfoil during air polishing.

3.4.3 Group III

The same procedures as for group II were carried out for this group, except that the air polishing was performed with glycine instead of sodium bicarbonate.

3.5 Biopsy procedure

Local anesthesia was applied remotely from the biopsy site in order to prevent any tissue damage from the injection. An internal bevel incision 2–3 mm paracrestally and a sulcular incision were made upon which a mucoperiosteal flap was reflected.

Figure 3.

Soft tissue analysis photomicrographs – histologic sections of biopsied tissues showing medium injury, i.e., superficial layers of the epithelium removed and basal membrane partially damaged.

3.6 Microscopic analysis

Microscopic analysis was carried out in the in vitro section of the soft tissue analysis. The biopsied tissues were prepared for histological sectioning by Gram staining and viewed under a light microscope with a 10 $\times$ zoom. Histological scoring of gingival tissue injury was done with respect to microscopically detectable tissue changes. (Fig. 3). Scoring was done using a 1–4 scale, based on the Petersilk G.J. method [18], with (1) No injury: undamaged epithelium and connective tissue; (2) Minor injury: disruption of superficial epithelial layers, undamaged basal membrane; (3) Medium injury: superficial layers of the epithelium removed, basal membrane partially damaged; (4) Severe injury: epithelium and basal membrane completely removed, connective tissue exposed.

4. Results

This study compared the values of surface changes in enamel and cementum as $R_{A}$ after SRP, SRP and SBAP and SRP and GPAP, followed by group comparisons (Table 1). Table 2 shows the mean and standard deviation of AFM values by site and by intervention. The mean AFM value was 108.5 and 144.7, 102.7 and 81.7, 95.6 and 7.4 respectively at the crown and at the root for the SRP, SRP and SBAP and SRP and GPAP interventions. The high standard deviation indicates there was high variation in the data. The minimum value was 16.6 and the maximum value was 262.7 at the crown site. Similarly, other intervention groups also had a large variation. Since the data was widely distributed, it was transformed to a logarithmic scale. The results of the two-way ANOVA infers that the ‘interaction effect, Site*intervention’ was non-significant, i.e., it also infers that the effect of all three interventions was similar for the two sites (crown and root) (Table 2).

Table 1
Atomic force microscopy values after SRP, SRP and sodium bicarbonate polishing, and SRP and glycine polishing

S. no.	Crown 1		Crown 2		Root 1		Root 2
SRP
1	26	.9	143	.3	97	.6	156	.2
2	110	.2	57	74	.5	174	.6
3	81	.7	79	82	.2	353	.2
4	38	.3	200	43	.6	171	.9
5	280	.6	69	.3	151	.3	132	.8
6	13	.7	102	.7	179	.4	119	.8
SRP and sodium bicarbonate polishing
1	217	25	.7	46	.I	59
2	47	.7	103	.3	45	.8	97	.7
3	262	.7	197	.5	76	.6	114
4	46	.1	37	.1	174	.8	101	.3
5	69	.8	16	.6	48	.9	67	.3
6	83	.7	125	.9	52	.7	59	.4
SRP and glycine polishing
1	93	.2	92	.3	98	.7	91	.3
2	101	.2	92	.7	93	.4	99	.5
3	89	.9	99	.1	98	.5	95	.7
4	97	.1	78	.2	89	88	.1
5	99	.7	98	.7	93	.2	93	.5
6	109	.1	96	.6	94	.6	78	.1

SRPs: Scaling and root planing.

Table 2

Mean and standard deviation of atomic force microscopy values by site and intervention

Intervention	Crown		Root		Two-way ANOVA test result ${}^{\#}$			LSD multiple
	Mean	SD	Mean	SD	Source	$F$ -value	$P$ -value	comparison test result
SRP	108.5	71.7	144.7	78.9	Site	0.595	0.443	SRP $>$ Sodium
Sodium bicarbonate	102.7	81.7	78.6	38.0	Intervention	3.196	0.047
$+$ SRP
Glycine $+$ SRP	95.6	7.4	92.8	5.8	Site*intervention	1.067	0.350

# Two-way ANOVA test has been applied for the logarithmic values. SRP: scaling and root planing.

The ‘main effect-site’ $p$ -value infers that the mean AFM values were equal for the two sites (crown and root) irrespective of the intervention received. The ‘main effect-intervention’ $p$ -value infers that the three interventions were statistically different at a 5% level of significance. To know which of the three groups differ, an LSD multiple comparison test was performed. The result infers that the AFM values were slightly higher in the SRP intervention group than the sodium intervention group. However, the glycine intervention group was not statistically different compared to the SRP intervention and sodium intervention group (Fig. 4). Soft tissue microscopic analysis showed that the minimum roughness was associated with glycine (Table 3).

Table 3

Mean value roughness score of various groups based on light microscopic analysis of histological sections

S. no. of patient	Group 1	Group 2	Group 3
1	4	3	2
2	3	2	1
3	3	3	2
4	4	3	1
5	3	1	1
6	4	3	2
Total	21	15	9
Mean value of roughness score	3.5	2.5	1.5

Figure 4.

Mean atomic force microscopy values site wise and intervention wise.

5. Discussion

A variety of prophylactic materials are used as part of routine procedures in the dental office for the removal of stains and calculus. Conventional methods such as rotating cups, brush cones and abrasive pastes which effectively remove all types of exogenous accumulations from the enamel surface, may cause undue abrasion or scratching of the enamel, dentin or cementum [11, 12, 13, 14]. As an alternative, air abrasion systems function by the abrasive action of a slurry of polishing powder ejected at high pressure through a nozzle surrounded by a cylindrical jet of water.

Various air polishing agents, such as sodium bicarbonate (baking soda), aluminum hydroxide, calcium carbonate, calcium phosphosilicate, bio active glass, glycine, are as efficient as a prophy cup and pumice on smooth surfaces and effectively remove stains [14, 15, 16, 17, 18, 19, 20, 21, 22] In the 1980–90s, sodium bicarbonate powder was most commonly used for air polishing to remove calculus and stains. However, it causes severe root damage with short application times during maintenance therapy and the cautious use of bicarbonates in patients on restricted sodium diets, respiratory or renal diseases and infectious diseases is recommended, which limits its use.

For efficient plaque removal, a low abrasive glycine is used for subgingival areas. In recent years, use of air polishing with glycine powder has been investigated with several in vitro and in vivo studies. Overall, these studies were consistent in indicating the clinical efficacy and low abrasive nature of glycine powder when sprayed on different dental and gingival structures [17].

The effect of polishing powders has been analyzed for more than three decades, and low abrasive powders have proved to be effective. With advances in nanotechnology, the use of AFM applications in dentistry is now popular as it allows for a high resolution, direct quantitative characterization of the surface roughness [12, 23]. In comparative studies of glycine and bicarbonate, glycine powder for 5 seconds was associated with the lower surface damage than observed with bicarbonate, under AFM [24]. Those powders are routinely used in restorative, orthodontic, and prosthetic dentistry for surface finish ad polishing [7, 15, 25, 26, 27, 28].

Regarding polishing devices, the routine use of hand pieces, rubber cups and bristle brushes have been used along with polishing powders or prophylaxis pastes [29]. Currently, a trend towards the use of polishing devices attracts for plaque removal even in inaccessible areas.

Supra- and subgingival biofilm removal is the prime objective of initial periodontal therapy and periodontal maintenance therapy (PMT). For mechanical debridement, hand instruments, sonic or ultrasonic scalers are used. The use of these instruments is technically demanding, and if debridement is performed periodically, tooth substance loss may occur with time [2, 18, 19]. Therefore, the use of air-polishing techniques may simplify periodic subgingival instrumentation and may be an alternative to the conventional techniques of biofilm removal. The air-abrasive system uses an abrasive powder introduced into a stream of compressed air to clean or polish a surface by removing deposits attached to it.

Regarding the efficacy of the system, it depends on various factors such as velocity of water and abrasive particles, distance between the nozzle tip and the tooth surface being treated, shape and size of abrasive particles [7, 11], duration of air polishing [30] and the amount of powder in the powder chamber. With this system, surface changes may be smooth or rough and this determines the plaque formation.

Microscopic surface details can be recorded with different instruments (profilometers, confocal microscopes, scanning electron microscopes, tunneling microscopes, but quantitative measurements at nano scale with AFM is the gold standard. It has the advantages of high resolution, nano measurements at particular point with high magnification and simple implementation, which is why it was used in our in vitro study. To define surface topography of the sample, R ${}_{a}$ the mean arithmetic roughness was measured, which shows that mean surface roughness was least with glycine. The two-way ANOVA results from the crown and root showed that the effect of all three interventions SRP, SBAP and GPAP was similar for the two sites (crown and root) and it was not statistically significant. The p value of AFM was slightly higher in the SRP group (0.443) when compared to the SBAP group (0.350). However, GPAP was not statistically different between the SRP intervention and SBAP group, which indicates that the three interventions were statistically different at a 5% level of significance, which correlates with previous studies.

The evaluations of the effects of air polishing on the human gingiva were based on clinical examination and localized soft tissue trauma as revealed by SEM inspection in earlier studies [31, 32]. The immediate effect of air-polishing devices on the histology of the gingiva was also evaluated [33]. On gingival exposure to air-polishing slurry it caused epithelial erosion, which positively correlated with instrumentation time and design principles of the applied air-polishing devices. Petersilka et al. [18] indicated that GPAP results in less gingival erosion than SBAP or hand instrumentation, further supporting the safety of the glycine air polishing technique. After 14 days of evaluation, the healing was similar with SBAP and GPAP. The present study, conducted with histological sections, agreed with this previous study [18]. It was determined from our study that SBAP and GPAP were comparatively better than hand instrumentation or ultrasonic devices. When comparing SBAP and GPAP, the latter was found superior to subgingival scaling or other polishing aids. In vitro analysis using AFM showed that surface roughness by SRP was slightly higher than SBAP, which indicates more surface roughness would result and more plaque would be retained, which could lead to increased colony forming units of microorganisms. Comparing the mean AFM values of the SRP (108.5-crown and 144.7-root), SBAP (102.7-crown and 78.6-root) and GPAP (95.6-crown and 92.8-root) groups no statistical differences were found. The histological soft tissue analysis indicated less soft tissue abrasion with GPAP than with SBAP and SRP.

5.1 Limitations

One limitation of this study was the small sample size used. Further studies with larger samples sizes should be conducted. Another limitation, which is inherent in any in vitro study is that the results cannot be generalized completely to in vivo conditions. Hence future clinical trials with other abrasive agents using air powder polishing systems could be tested. It is recommended that long-term in vivo studies are carried out to evaluate the microbial accumulation on the rough tooth surface areas.

6. Conclusions

It can be concluded that SBAP and GPAP were better than hand instrumentation, as proved in our study by AFM and histological section analysis. GPAP is safer, superior, and more comfortable. It can also be applied subgingivally for biofilm removal. The frequent use of mechanical instrumentation may produce surface alterations during periodontal therapy. As an alternative, air polishing devices can be used for plaque removal as it is easier and causes less damage.

Footnotes

Conflict of interest

None to report.

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Comparison of the effect of sodium bicarbonate and glycine air polishing systems on tooth surface roughness: An atomic force microscopic analysis

Abstract

BACKGROUND:

OBJECTIVE:

METHODS:

RESULTS:

CONCLUSION:

Keywords

1. Introduction

2. Methods

3. In vitro phase – Hard tissue analysis

3.1 Tooth selection and preparation

3.3 AFM measurements

3.4.1 Group I

3.4.2 Group II

3.4.3 Group III

3.5 Biopsy procedure

4. Results

Table 1 Atomic force microscopy values after SRP, SRP and sodium bicarbonate polishing, and SRP and glycine polishing

5.1 Limitations

6. Conclusions

Footnotes

Conflict of interest

References

Table 1
Atomic force microscopy values after SRP, SRP and sodium bicarbonate polishing, and SRP and glycine polishing