Synthesis and toxicity evaluation of a spray consisting of silver nanoparticles, ethylenediaminetetraacetic acid, methylsulfonylmethane, and xylitol on vero cell line

Abstract

Introduction: Dental plaque is a biofilm or an accumulation of bacteria that grows in the internal surfaces of the mouth and can be observed as a white to pale yellow layer over or between teeth. The continuous formation and accumulation of dental plaque lead to oral diseases. As a result, it is necessary to prevent the aggregation of dental plaque and clean it daily. Recently, ethylenediaminetetraacetic acid (EDTA) has been utilized in toothpaste to prevent plaque formation as an EDTA-transporting enhancer, methylsulfonylmethane (MSM) can effectively increase its local effect. Xylitol decreases the Streptococcus mutans count by changing the metabolic pathways. Aims: In the present study, we synthesized a solution containing silver nanoparticles, EDTA, MSM, and xylitol and evaluated its toxicity on the Vero cell line through the MTT assay. Materials and Methods: To produce silver nanoparticles, we dissolved silver nitrate in sodium citrate. Then we used a solution of distilled water and polyvinylpyrrolidone, which resulted in the encapsulation and stabilization of silver nanoparticles, and the solution was made by mixing other ingredients. We evaluated the cytotoxicity of this spray using the Vero cell line. We cultured the cells in the 10% FBS-containing RPMI culture medium. For performing the cytotoxicity test, we put 10,000 cells in each well of the 96-well plate, and the next day added the synthesized solution to each well at dilutions 0.05%, 0.1%, 0.2%, 0.5%, 1%, 2%, 5%, and 10%. As the control group, we used 4 wells containing live Vero cells without adding the solution. After 24, 48, and 72 h, we added the MTT solution to each well and incubated the plates at 37°C for 4 h. Finally, we evaluated the rate of living cells by reading the absorbance with an ELISA device at 570 nm. Results and Discussion: We used the Mann–Whitney nonparametric test to evaluate the cytotoxicity of different concentrations of the synthesized spray solution and compare the cell viability rate of groups with the controls in various periods. According to the cytotoxicity results of different concentrations of the spray solution on the Vero cell line, there was no significant difference in cytotoxicity between the 0.05%, 0.1%, 0.2%, and 0.5%, and control groups at 24, 48, and 72 h (P > 0.05). No significant difference existed in cytotoxicity between 1% and 2% concentrations and the controls 24 h after exposure; this became significant after 48 and 72 h (P = 0.014). However, a significant difference existed in cytotoxicity between 5% concentration and the controls 24, 48, and 72 h after exposure (P = 0.014). The CC50 of the spray solution was calculated at 3.51%. Conclusion: The findings of this study showed that the synthesized solution is nontoxic; therefore, this spray solution can be used safely as an oral mouthwash and spray.

Keywords

• cell toxicity
• ethylenediaminetetraacetic acid
• methylsulfonylmethane
• mtt assay
• nanosilver
• vero cells
• xylitol