An in vitro study was conducted from September 2016-October 2016 on dentin samples obtained from extracted human maxillary and mandibular both upper and lower premolars. The study was conducted at Government Dental College and Research Institute Bangalore and Indian Institute of Technology Bengaluru, Karnataka, India. Teeth with untreated or treated caries, hypoplasia, wear and trauma were excluded from the study.

A total of 90 dentin samples were obtained from 45 premolars which were sectioned mesio-distally using diamond disc to obtain two samples per tooth. The dentin specimen of 5*4*4 mm was obtained using airotar and long straight bur. All the specimens were treated with 6% citric acid for two minutes to facilitate removal of the smear layer and to disocclude the dentinal tubules. These treated specimens were washed thoroughly with distilled water to remove any remaining acid or debris present. The specimens were then randomly assigned to one of the following three groups [Table/Fig-1].

The intervention was manual application of toothpaste (rubbing of toothpaste on dentin with finger) and immersion of specimens in mouthwash for Group 1 and 2 respectively whereas Group 3 received no intervention (control). Samples in Group 1 and 2 after the intervention were placed in distilled water. At the end of 3 days, 7 days and 14 days, ten samples each from all group (five from each subgroups) were subjected to SEM to detect tubular occlusion. Equal samples were taken from subgroups.

Two blinded evaluators were trained and calibrated (inter and intra examiner reliability scores being 0.83 and 0.85 respectively) for the evaluation of dentin occlusion based on criteria in the previous literature [9].

Type 0: Open dentinal tubules without occlusion i.e., acid treated specimens without desensitising agent application.

Type 1: Partial dentinal tubule occlusion < 25% of dentinal tubule orifice.

Type 2: Partial dentinal tubule occlusion of > 25% up to75% of dentinal tubule orifice.

Type 3: Nearly complete dentinal tubule occlusion of >75% of dentinal tubule orifice.

At the time of analysis, Type 2 and Type 3 were considered in the same category as “partial dentinal tubule occlusion”.

Study armamentarium included gloves, mouth mask, diamond disc, straight airotar, burs, extracted teeth, citric acid, potassium nitrate toothpaste and mouthwash, dapen dish and distilled water. SEM (Zeiss sigma VP, Zeiss Oberkochen, Germany) was used for the analysis.

Data obtained was entered in a MS Excel sheet. The descriptive statistics were computed with the Statistical Package of Social Sciences (SPSS) version 22.0 software. Chi-square test was applied to assess the significant difference between the groups. The p-value was considered as significant when less than 0.05 (confidence interval of 95%).

The present study evaluated the dentinal tubule occluding effect of potassium nitrate applied as toothpaste and mouthwash, once/ twice daily for 3, 7 and 14 days.

SEM images of different groups at the end of each period are reproduced in [Table/Fig-2-6]. At the end of third day, dentinal tubules were partially occluded in Group 1 {subgroups a and group 2 (subgroup b)} [Table/Fig-7]. There was a statistically significant difference inthe tubular occlusion between the groups at the end of 3^rd day (p<0.001).

[Table/Fig-2]:

Control group at the end of 3^rd, 7^th and 14^th day.

[Table/Fig-3]:

Group 1a at the end of 3^rd, 7^th, 14^th days.

[Table/Fig-4]:

Group 1b at the end of 3^rd, 7^th, 14^th day.

[Table/Fig-5]:

Group 2a at the end of 3^rd, 7^th, 14^th day.

[Table/Fig-6]:

Group 2b at the end of 3^rd, 7^th, 14^th day.

At the end of 7^th day, dentinal tubules were either partially occluded (subgroup a) or both partially occluded and nearly complete occluded (subgroup b) in Group 1 whereas dentinal tubules were partially occluded in Group 2 (subgroups a and b) [Table/Fig-8]. A statistically significant difference was observed in the tubular occlusion between the groups at the end of 7^th day (p<0.001).

At the end of 14^th day, dentinal tubules were both partially occluded and nearly complete occluded (subgroup a) in Group 1 and nearly complete occluded (subgroup b) in group 1, whereas dentinal tubules were both partially occluded and nearly complete occluded (subgroup a and b) in Group 2 [Table/Fig-9] and the difference was found to be statistically significant (p<0.001).

However, the proportion of samples which were nearly complete occluded in the decreasing order were Group 1 (subgroup b), group 1 (subgroup a), Group 2 (subgroup b) and Group 2 (subgroup a).

The present study evaluated and compared the in vitro efficacy of potassium nitrate as a dentin tubule occluding agent in toothpaste and mouthwash form, applied once/ twice daily for 3, 7 and 14 days. Not many in vitro studies have been reported in literature on efficacy of potassium nitrate as a dentin occluding agent. Therefore comparisons are made wherever feasible.

An animal study on rats has demonstrated efficacy of potassium nitrate in tubular occlusion [10]. An in vivo demonstrated that the use of toothpaste containing 5% potassium nitrate on hypersensitive teeth was significantly more effective in immediate pain reduction than using placebo toothpaste [11]. Studies have found potassium nitrate to be inferior to calcium sodium phospho silicate [12] and Novamin [13]. However, studies have reported improved clinical effectiveness when potassium nitrate was combined with other agents like sodium fluoride and cetylpyridinium chloride [14] as well as with sodium monofluorophosphate, nano-hydroxyapatite, antioxidants phloretin, ferulic acid and silymarin [15]. In contrast to our results, an in vivo study demonstrated equal effectiveness of potassium nitrate toothpaste and mouthwash in reducing sensitivity when applied for four weeks [5].

Although, the effect of potassium nitrate mouthwash was lower as compared to toothpaste, tubular occlusion was seen in group 2a and 2b, with twice daily application showing better results. This is in line with another study where it was reported that potassium nitrate mouthwash is effective in alleviating dentinal sensitivity [16].

The result of our study showed that potassium nitrate in toothpaste form is more efficacious than mouthwash form in occluding the dentin tubules. Similarly, it was more efficacious when applied twice daily in both the groups. Maximum efficacy was observed at 14^th day for toothpaste applied twice daily. The better tubular occluding effect of toothpaste form and the higher concentration of active ingredient (potassium nitrate) could be partly attributed to the better retention of the material and occlusion produced by abrasive components of toothpaste.

However, the study has some limitations. This study employed in vitro design, hence limiting its ability to mimic oral health conditions. Secondly, after application, all the specimens were placed in distilled water. Hence, the impact of oral environment post application could not be reproduced in this study. Also, its efficacy when compared to other agents could not be as certained which was beyond the scope of the study. Since dentinal hypersensitivity is subjective in nature, the results need to be validated against patient’s perception of sensitivity quality of life. Hence, further in vivo studies are required on effectiveness of potassium nitrate in different forms, frequency and duration in comparison with other agents.

The present in vitro study demonstrated maximum tubular occluding effects of potassium nitrate when applied in toothpaste form, twice daily for a period of two weeks. Further long term in vivo studies are suggested.