| Dentin hypersensitivity (DH) is a common and frequently-occurring disease, and the morbidity increases with age, which raises the interest of researchers. Hydroxyapatite (HA), an important natural mineral substance of the human body hard tissues, has good biocompatibility and bioactivity, and consists of the enamel by98%. At present, it is evidenced that dentifrice containing HA has the role of anti-sensitivity which mixes with HA as the active ingredient resulting from its characteristic of homogeneous phase adsorption. However, this fild is still in the state of full function connection, dentifrice containing HA has not been widely used, related to the requirement of technology and high cost. With the developing of nanotechnology, nano-material has been concerned for the outstanding characteristics of nano-particles. We wonder if dentifrice containing nano-HA has better effect on plugging the dentinal tubules for anti-sensitivity than that containing common sized particles has. On the other hand, a lot of studies have found that HA has a good adsorption capacity to heavy metal ions, and nano-HA has the better effect. But there has not yet been reported about whether HA mixed in dentifrice still has the adsorption effect to heavy metals in aqueous solution. It is still not clear that if there are differences among the adsorption capacity of different particle sizes of HA mixed in dentifrice and what is the best. If HA mixed in dentifrice has the adsorption capacity on the metal ions, how about the stability of this adsorption force?ObjectiveTo unify and standardize ripping method of dentin disk, a comparative study of different concentrations of citric acid solution on removing the smear layer and exposing the dentinal tubules ability at different times was designed, so the best concentration of citric acid and etch time on preparing dentin hypersensitive model would be achieved, and a reliable in vitro model for studying dentin hypersensitivity would be obtained. By simulating an oral environment, two preliminary comparative study of effect of dentifrice containing HA with different particle sizes on dentinal tubule occlusion were performed quantitatively and qualitatively; Measuring the remineralization situation after blushed with dentifrice containing HA; The stability of dentinal tubule occlusion was investigated by brushing on the dentin surface.To explore and compare the capacity of dentifrice containing HA with different particle sizes on aqueous Cr (Ⅵ) sorption, as well as the stability of the sorption capacity.Methods(1) Thirty premolars (removed for orthodontic treatment and the crown were intact) were selected, removing the occlusal enamel layer under cooling water. Parallel to the occlusal surface, fine carborundum needle cut at about1.5mm under the dentino-enamel junction of the premolar crown, and dentin pieces with thickness of1.5mm were obtained. Each dentin disc surface was wet polished using silicone carbide paper grit600,1000,1200, and2000to form a uniform, smooth and plat surface (away from the canal side). Then the samples were examined under a microscope for confirming that they had been polished, without surface scratches, consistent transmittance. The polished samples were cleaned ultrasonically to remove impurities in de-ionized water for10min, three times in a row, and then rinsed three times using a large quantity of de-ionized water. So the dentin disk simulating dentin-sensitive model was made for the next experiment. Different concentrations of citric acid (5%,10%,15%) was used to etch the dentin discs at different times (30,60,120,180s), removing the etching solution, cleaning ultrasonically in de-ionized water and drying naturally in the air. The dentin disk surfaces were sprayed gold, and observed by scanning electron microscopy. The Image-pro plus6.0was used to measure the diameter of the dentinal tubules. The effects of opening dentinal tubules and removing the smear of dentin disk surface were analyzed comparatively among different concentrations of citric acid solution (5%,10%,15%) etched the dentin samples at different times (30,60,120,180s) respectively by using SPSS16.0statistical analysis software.(2) Seventy dentin disks of70molars and70premolars were prepared in accordance with the first experiment, and randomly selected20dentin disks (10molars and10premolars) as untreated group for EDS examination. The others were divided into6groups:Control group without using any experimental treatment, Negative control group treating with dentifrice not containing HA, Positive control group treating with Colgate sensitive pro-relief dentifrice, Experimental group1treating with dentifrice containing3%80nm HA, Experimental group2treating with dentifrice containing3%300nm HA, and Experimental group3treating with dentifrice containing3%600nm HA. Each dentin disk was brushed with the same amount of dentifrice (0.2g), washed with distilled water for2min, the guinea pigs were removed under distilled water for30s, then stored in artificial saliva (pH=6.8) at37℃water bath, twice everyday. Seven days later, the dentin disk was divided into two parts, and half of one was detected by the SEM and EDS, blushed with distilled water for30s before detecting; the other one was brushed with distilled water for2min, washed with distilled water for30s, and stored in artificial saliva (pH=6.8) at37℃water bath, twice everyday. Seven days later, the samples were dried in air and observed by SEM.(3) One ml dentifrice solution (dentifrice containing80nm HA, dentifrice containing300nm HA, dentifrice containing600nm HA, blank dentifrice), with the concentration of0.025g/100ml,0.05g/100ml,0.1g/100ml,0.2g/100ml,0.3g/100ml,0.4g/100ml,0.5g/100ml,0.6g/100ml,0.7g/100ml,0.8g/100ml,0.9g/100ml1.0g/100ml, was added to50ml potassium dichromate solution with the initial concentration of100μl/ml respectively. When the reaction reached equilibrium, the mixed solution was centrifugated at3000r/min for10min at the temperature of25℃. The residual heavy metal ion concentration of the supernatant was measured by diphenylcarbon phthalocyanine hydrazine method, and the removal rate was calculated. The precipitate of the solution was dried at120℃, and the elements constitute was detected by EDS. The above procedure was repeated, the mixed solution was placed at room temperature for14d,28d, then the residual heavy metal ion concentration and the elements constitute were measured again.Results(1) After etching30s,60s,120s, or180s with5%citric acid solution, as well as30s, or60s with10%or15%citric acid solution, the ability of removing the smear layer is limited, the dentinal tubules are not opened completely, and there are still some dentinal tubules covered by the smear layer in different degrees. The smear layer can be removed from dentinal tubules, and the dentinal tubules are exposed completely after the dentin discs were etched with10%,15%citric acid solution for120s, or180s. Statistical analysis demonstrated, there are significant statistically differences between different etching time (F=52.948, P=0.000); Also, There are significant statistically differences between different concentration (F=15.765, P=0.000); however, there is no interaction effect between different etching time and concentrations (F=4.292, P=0.051). Therefor, only the main effect of time and concentration should be analyzed. Using two-sample t-test for pairwise comparisons of10%citric acid for120S、180s and15%citric acid for120s、180s four groups, we found that there were significant differences between15%citric acid for120s and10%citric acid for120s (t=4.860, P=0.001),15%citric acid for180s and10%citric acid for180s (t=5.502, P=0.001), it is mean that the dentinal tubules diameter of15%concentration group are greater than the10%concentration group; There is no significant difference between10%citric acid for120s、180s (7=1.769, P=0.107).(2) After treatment by dentifrice for7d, the plugging rate of the dentinal tubules in the negative control group is lower than other treated groups (molar group:χ2=98.873, P=0.000; premolar group:χ2=96.644, P=0.000), and the plugging rate of Colgate sensitive pro-relief dentifrice group is the highest. While the plugging rates decrease progressively with the turn of dentifrice containing80,300,600nm HA, the80nm group get the best effect:the diameter of dentinal tubules is smaller, nearly all the dentinal tubules are blocked by the dentifrice materials, the materials particles are tiny, tightly occlusion, and the boundary between the material and the inter-tubular dentin is not clear. Then after cleaned by distilled water for7d, similar results of plugging rate of dentinal tubules are obtained (molar group:χ2=85.622, P=0.000; premolar group:χ2=91.881, P=0.000), except600nm group have better blocking effect than300nm group. Energy spectrum analysis showed that there are significant statistically differences among the atomic percentage of Ca and P on the surface of dentinal tubules in each experimental group of the molar group (comparison of Ca element: χ2=47.883,P=0.000; comparison of P element:χ2=48.184, P=0.000). The percentage of Ca and P element among the molars groups is:80nm group>600nm group>300 nm group> negative control group> blank control group> untreated group. A similar results of the percentage of Ca and P element among the molars groups is found among premolars groups (comparison of Ca element:χ2=47.922, P=0.000; comparison of P element:χ2=49.445, P=0.000).(3) The concentration of aqueous Cr (VI) decreases progressively as the concentra-tion of the dentifrice adding to the solution increases. The chromium ion adsorption rate of the dentifrice containing600nm HA group is the highest for52.36%; dentifrice containing80nm HA group followed by49.29%; dentifrice containing300nm HA group by44.72%; and the blank control group by33.43%. That is to say that the adsorption capacity of per gram80nm,300nm, and600nm HA dentifrice on the chromium ion are246.45mg,223.6mg, and261.8mg. Chromium ion can be detected in the precipitate, further confirming the chromium in the chromium ion solution is adsorbed by dentifrice containing HA. The aqueous Cr (VI) sorption of dentifrice containing80nm、600nm HA group appears stable after the mixed solution was placed at room temperature for14d and28d, while dentifrice containing300nm HA shows desorption phenomenon in the region of0.025-0.7g/100ml, and the adsorption rate of blank dentifrice increases over time in the concentration region of0.025～0.5g/100ml. It is inferred that dentifrice containing80nm or600nm HA may be a good adsorbent of chromium ion.Conclusions(1)10%citric acid for120s is the best etching condition for removing the smear layer of the dentin discs and acquiring a reliable in vitro model for dentin hypersensitivity study.(2) Dentifrice containing HA is, especially80nm HA dentifrice, of good dentinal tubule occlusion and surface mineralization effect.(3) Dentifrice containing HA is, especially600nm HA dentifrice, a good adsorbent of aqueous Cr (Ⅵ). |
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