'Green' silver nanoparticles combined with tyrosol as potential oral antimicrobial therapy.

OBJECTIVES: This study aimed to evaluate silver nanoparticles (AgNPs) obtained by a 'green' route associated or not to tyrosol (TYR) against Streptococcus mutans and Candida albicans in planktonic and biofilms states. METHODS: AgNPs were obtained by a 'green' route using pomegranate extract. The minimum inhibitory concentration (MIC) against S. mutans and C. albicans was determined for AgNPs and TYR combined and alone, and fractional inhibitory concentration index (FICI) was calculated. Single biofilms of C. albicans and S. mutans were cultivated for 24 h and then treated with drugs alone or in combination for 24 h. RESULTS: AgNPs and TYR were effective against C. albicans and S. mutans considering planktonic cells alone and combined. The MIC values obtained for C. albicans was 312.5 µg/mL (AgNPs) and 50 mM (TYR) and for S. mutans was 78.1 µg/mL (AgNPs) and 90 mM (TYR). The combination of these antimicrobial agents was also effective against both microorganisms: 2.44 µg/mL/0.08 mM (AgNPs/TYR) for C. albicans and 39.05 µg/mL /1.25 mM (AgNPs/TYR) for S. mutans. However, synergism was observed only for C. albicans (FICI 0.008). When biofilm was evaluated, a reduction of 4.62 log10 was observed for S. mutans biofilm cells treated with AgNPs (p < 0.05, Tukey test). However, the addition of TYR to AgNPs did not improve their action against biofilm cells (p > 0.05). AgNPs combined with TYR demonstrated a synergistic effect against C. albicans biofilms. CONCLUSIONS: These findings suggest the potential use of AgNPs with or without TYR against C. albicans and S. mutans, important oral pathogens. CLINICAL SIGNIFICANCE: AgNPs obtained by a 'green' route combined or not with TYR can be an alternative to develop several types of oral antimicrobial therapies and biomaterials.

Buffering Capacity and Effects of Sodium Hexametaphosphate Nanoparticles and Fluoride on the Inorganic Components of Cariogenic-Related Biofilms In Vitro.

Despite the remarkable effects of sodium hexametaphosphate nanoparticles (HMPnano) on dental enamel de-/re-mineralization processes, information on the effects of these nanoparticles on biofilms is scarce. This study assessed the effects of HMPnano, with or without fluoride (F), on the inorganic components and pH of Streptococcus mutans and Candida albicans dual-species biofilms. Solutions containing conventional/micro-sized HMP (HMPmicro) or HMPnano were prepared at 0.5% and 1%, with or without 1100 ppm F. A 1100 ppm F solution and pure artificial saliva were tested as positive and negative controls, respectively. The biofilms were treated three times and had their pH analyzed, and the concentrations of F, calcium, phosphorus, and HMP in the biofilm biomass and fluid were determined. In another set of experiments, after the last treatment, the biofilms were exposed to a 20% sucrose solution, and the biofilm pH and inorganic components were evaluated. The 1% HMPnano solution with F led to the highest biofilm pH, even after exposure to sucrose. The 1% HMPnano solution without F led to significantly higher phosphorus concentrations in comparison to all other groups. It can be concluded that 1% HMPnano and F influenced the biofilm pH, besides affecting most of the inorganic components of the dual-species biofilms.

Fluoride and trimetaphosphate association as a novel approach for remineralization and antiproteolytic activity in dentin tissue.

OBJECTIVE: The study evaluated the effect of solutions containing fluoride (F) and/or sodium trimetaphosphate (TMP) and F/TMP on the inhibition of MMP-2 and MMP-9, and on dentin remineralization in vitro. DESIGN: Bovine root dentin blocks were prepared, and caries-like lesions were induced in two thirds of the surface. Blocks were then randomly divided into 13 groups/solutions (n = 10): Placebo; 0.3 %, 1 % and 3 % NaOH-hydrolyzed TMP; 0.3 %, 1 % and 3 % TMP; 250, 500 and 1100 ppm F; 250 ppm F + 0.3 % TMP; 500 ppm F + 1 % TMP and 1100 ppm F + 3 % TMP. One third of each specimen was treated with the respective solutions in pH-cycling. The mineral concentration (gHAp × cm-3 × µm) was determined by computed X-ray microtomography, and data submitted to ANOVA and Student-Newman-Keuls' test (p < 0.05). The ability of the solutions to inhibit MMP-2 and MMP-9 activity was assessed by zymography. RESULTS: F/TMP association led to less mineral loss in the deeper region of the lesion and reduced the depth of lesions when compared to its counterpart without TMP (p < 0.001). 3 % TMP (hydrolyzed or not), 500 ppm F and 1100 ppm F completely inhibited MMP-2 activity, while for MMP-9 such effects were only achieved by treatment with 1100 ppm F + 3 % TMP. CONCLUSION: Treatment with 1100 ppm F + 3 % TMP fully inhibits the gelatinolytic activity of MMPs-2 and - 9 and shows greater remineralizing capacity in artificial caries lesions in dentin. However, hydrolyzing TMP does not improve its anti-proteolytic activity and its remineralizing capacity.

In vitro Evaluation of Surface Free Energy of Dentin after Treatment with Sodium Trimetaphosphate Associated or Not with Fluoride, Exposed or Not to Calcium.

It has been stated that sodium trimetaphosphate (TMP) promotes a more anionic dentin surface inducing greater calcium (Ca) and phosphate precipitation. The aim of the present study was to evaluate in vitro the surface free energy (γs) of dentin after treatment with TMP associated or not with fluoride (F), exposed or not to Ca, as well as the adsorption of TMP, F, and Ca by dentin. Bovine dentin blocks (n = 12 blocks/group) were treated with solutions containing TMP at 0, 1, 3, or 9 (w/v) followed or not by the application of Ca. These solutions were or were not associated to 1,100 ppm F. F, Ca, and TMP were determined in the solutions before and after the treatment to calculate the adsorption by dentin. To analyze the γs of dentin, the apolar (γsLW), and polar (γsAB), components were determined by contact angle measurement. Data were submitted to 2-way ANOVA followed by the Student-Newman-Keuls test (p < 0.05). TMP reduces γs of dentin and increases electron donor sites (γs-). Higher values of γs- led to higher adsorption of Ca (p < 0.001). The F/TMP association did not change γs or γsLW and reduced the values of γs-, but the adsorption of Ca was higher. There was correlation between the adsorption of TMP and γs- (Pearson's r = 0.801; p < 0.001) and F (Pearson's r = 0.871, p < 0.001). It is possible to conclude that TMP increased γs- and Ca adsorption, and reduced γs. The association with F increased the adsorption of TMP without rising γs-; however, there was higher adsorption of Ca.

Effect of S. mutans combinations with bifidobacteria/lactobacilli on biofilm and enamel demineralization.

The present study evaluated the ability of Bifidobacterium and Lactobacillus species associated with streptococci to increase insoluble extracellular polysaccharide (EPS) production and initial caries lesion progression. Bovine enamel blocks (n = 190; 4 mm x 4 mm) were prepared, selected according to initial surface hardness (SH), and divided into two groups: a) double combinations: S. mutans with Bifidobacterium or Lactobacillus, and b) triple combinations: S. mutans and S. sobrinus with Bifidobacterium or Lactobacillus species. The blocks were exposed to the bacterial associations for 7 days. Subsequently, quantity of EPS from biofilms and caries lesion depth were determined by means of colorimetric and cross-sectional enamel hardness (ΔKHN) analysis. The data were submitted to one-way analysis of variance, followed by the Bonferroni test (p < 0.05). S. mutans with B. animalis or B. dentium produced a higher quantity of EPS; S. mutans + B. animalis led to the highest ∆KHN. S. mutans + S. sobrinus + B. longum induced greater EPS and ∆KHN values. In conclusion, associations of B. animalis and B. longum with streptococci promoted EPS production and caries lesion progression.

Effect of S. mutans combinations with bifidobacteria/lactobacilli on biofilm and enamel demineralization

Abstract The present study evaluated the ability of Bifidobacterium and Lactobacillus species associated with streptococci to increase insoluble extracellular polysaccharide (EPS) production and initial caries lesion progression. Bovine enamel blocks (n = 190; 4 mm x 4 mm) were prepared, selected according to initial surface hardness (SH), and divided into two groups: a) double combinations: S. mutans with Bifidobacterium or Lactobacillus, and b) triple combinations: S. mutans and S. sobrinus with Bifidobacterium or Lactobacillus species. The blocks were exposed to the bacterial associations for 7 days. Subsequently, quantity of EPS from biofilms and caries lesion depth were determined by means of colorimetric and cross-sectional enamel hardness (ΔKHN) analysis. The data were submitted to one-way analysis of variance, followed by the Bonferroni test (p < 0.05). S. mutans with B. animalis or B. dentium produced a higher quantity of EPS; S. mutans + B. animalis led to the highest ∆KHN. S. mutans + S. sobrinus + B. longum induced greater EPS and ∆KHN values. In conclusion, associations of B. animalis and B. longum with streptococci promoted EPS production and caries lesion progression.

Study of the activity of Punica granatum-mediated silver nanoparticles against Candida albicans and Candida glabrata, alone or in combination with azoles or polyenes.

The continuous emergence of Candida strains resistant to currently used antifungals demands the development of new alternatives that could reduce the burden of candidiasis. In this work silver nanoparticles synthesized using a green route are efficiently used, alone or in combination with fluconazole, amphotericin B or nystatine, to inhibit growth of C. albicans and C. glabrata oral clinical strains, including in strains showing resistance to fluconazole. A potent inhibitory effect over biofilm formation prompted by the two Candida species was also observed, including in mature biofilm cells. These results foster the use of phytotherapeutics as effective treatments in oral candidiasis.

Antimicrobial Activity of Compounds Containing Silver Nanoparticles and Calcium Glycerophosphate in Combination with Tyrosol.

Nanocomposites containing antimicrobial agents and calcium phosphates have been developed. Thus, this study assessed the effects of two compounds containing silver nanoparticles (AgNPs) and ß-calcium glycerophosphate (CaGP), associated or not with tyrosol (TYR), against planktonic cells and biofilms of Candida albicans and Streptococcus mutans. The nanocompounds were synthesized through chemical and 'green' processes and characterized by scanning electron microscopy. The minimum and fractional inhibitory concentrations of each compound were determined for planktonic cells. Next, 24-h single biofilms of C. albicans and S. mutans were treated for 24 h with the nanocompounds alone or in combination with TYR, and the antibiofilm effect was assessed through enumeration of colony forming units. Biofilm data were statistically examined using one-way ANOVA and the Kruskal-Wallis test (α = 0.05). The chemically synthesized nanocompound in combination with TYR demonstrated a synergistic effect against planktonic cells of C. albicans and S. mutans. For the nanocompound obtained through the 'green' route associated with TYR, a synergistic effect was observed only against C. albicans. For biofilms, only the combination obtained through the 'green' route + TYR demonstrated a synergistic effect against C. albicans. Our results may contribute to the development of oral care products containing AgNPs-CaGP and TYR to combat oral infections.

Resin-modified glass ionomer containing calcium glycerophosphate: physico-mechanical properties and enamel demineralization.

BACKGROUND: Sources of calcium and phosphate have been added to dental restorative materials to improve their anticaries effect. OBJECTIVE: This study evaluated the effect of adding calcium glycerophosphate (CaGP) to resin-modified glass ionomer cement (RMGIC) on the physico-mechanical properties, ion release, and enamel demineralization. Material and Methods: Specimens were fabricated for each experimental group: RMGIC without CaGP (Control), RMGIC with 1, 3 and 9% CaGP. To determine the release of fluoride (F), calcium (Ca) and phosphorus (P), six specimens were immersed in demineralization and remineralization solutions for 15 days. In another experimental trial, the following physico-mechanical properties were evaluated at time intervals of 1 and 7 days after fabrication: compressive strength (n=12), diametral tensile strength (n=12), surface hardness of material (n=6) and the degree of conversion of monomers (n=8). To study enamel demineralization, specimens (n=12) were attached to enamel blocks and submitted to pH-cycling. Subsequently, surface and cross-sectional hardness and the concentration of F, Ca and P in enamel were determined. RESULTS: The addition of CaGP to RMGIC led to higher mean release of F, Ca and P when compared with control (p<0.001). Mechanical properties were within the range of those of the ionomer cements after addition of 1% and 3% CaGP. The degree of conversion did not differ between groups at the 1st and the 7th day (p>0.439). The addition of 3% and 9% CaGP reduced mineral loss and increased F, Ca and P in the enamel when compared with control (p<0.05). CONCLUSION: The addition of 3% CaGP in RMGIC increased the release of F, P and Ca, reduced enamel demineralization, and maintained the physico-mechanical properties within the parameters for this material.

Ion release, antimicrobial and physio-mechanical properties of glass ionomer cement containing micro or nanosized hexametaphosphate, and their effect on enamel demineralization.

OBJECTIVES: To evaluate the effects of hexametaphosphate microparticles (mHMP) or nanoparticles (nHMP) incorporated in glass ionomer cement (GIC) on antimicrobial and physico-mechanical properties, fluoride (F) release, and enamel demineralization. MATERIAL AND METHODS: HMP solutions were obtained at concentrations of 1, 3, 6, 9, and 12%, for screening of antimicrobial activity. Next, mHMP or nHMP at 6, 9, and 12% were incorporated into a resin-modified GIC and the antibacterial activity was evaluated. The resistance to diametral tensile and compressive strength, surface hardness, and degree of monomer conversion as well as F and HMP releases of GICs were determined. Furthermore, specimens were attached to enamel blocks and submitted to pH-cycling, and mineral loss was determined. Parametric and non-parametric tests were performed, after checking data homoscedasticity (p < 0.05). RESULTS: HMP solutions at 6, 9, and 12% demonstrated the best antibacterial activity. GIC containing HMP showed better antibacterial effects at 9 and 12% for nHMP. Regarding F and HMP releases, the highest levels of release occurred for groups containing 9 and 12% nHMP. With the increase in HMP concentration, there was lower mineral loss. However, the incorporation of mHMP or nHMP in GIC reduced values of physico-mechanical properties when compared to the control GIC. CONCLUSIONS: nHMP improves antimicrobial activity and fluoride release, and decreases enamel demineralization, but reduces the physico-mechanical properties of GIC. CLINICAL RELEVANCE: The association of GIC/HMP could be an alternative material for patients at high risk for dental caries and could be indicated for low-stress regions or provisional restorations.

Resin-modified glass ionomer containing calcium glycerophosphate: physico-mechanical properties and enamel demineralization

Abstract Sources of calcium and phosphate have been added to dental restorative materials to improve their anticaries effect. Objective This study evaluated the effect of adding calcium glycerophosphate (CaGP) to resin-modified glass ionomer cement (RMGIC) on the physico-mechanical properties, ion release, and enamel demineralization. Material and Methods: Specimens were fabricated for each experimental group: RMGIC without CaGP (Control), RMGIC with 1, 3 and 9% CaGP. To determine the release of fluoride (F), calcium (Ca) and phosphorus (P), six specimens were immersed in demineralization and remineralization solutions for 15 days. In another experimental trial, the following physico-mechanical properties were evaluated at time intervals of 1 and 7 days after fabrication: compressive strength (n=12), diametral tensile strength (n=12), surface hardness of material (n=6) and the degree of conversion of monomers (n=8). To study enamel demineralization, specimens (n=12) were attached to enamel blocks and submitted to pH-cycling. Subsequently, surface and cross-sectional hardness and the concentration of F, Ca and P in enamel were determined. Results The addition of CaGP to RMGIC led to higher mean release of F, Ca and P when compared with control (p<0.001). Mechanical properties were within the range of those of the ionomer cements after addition of 1% and 3% CaGP. The degree of conversion did not differ between groups at the 1st and the 7th day (p>0.439). The addition of 3% and 9% CaGP reduced mineral loss and increased F, Ca and P in the enamel when compared with control (p<0.05). Conclusion The addition of 3% CaGP in RMGIC increased the release of F, P and Ca, reduced enamel demineralization, and maintained the physico-mechanical properties within the parameters for this material.

Nanosynthesis of Silver-Calcium Glycerophosphate: Promising Association against Oral Pathogens.

Nanobiomaterials combining remineralization and antimicrobial abilities would bring important benefits to control dental caries. This study aimed to produce nanocompounds containing calcium glycerophosphate (CaGP) and silver nanoparticles (AgNP) by varying the reducing agent of silver nitrate (sodium borohydride (B) or sodium citrate (C)), the concentration of silver (1% or 10%), and the CaGP forms (nano or commercial), and analyze its characterization and antimicrobial activity against ATCC Candida albicans (10231) and Streptococcus mutans (25175) by the microdilution method. Controls of AgNP were produced and silver ions (Ag⁺) were quantified in all of the samples. X-ray diffraction, UV-Vis, and scanning electron microscopy (SEM) analysis demonstrated AgNP associated with CaGP. Ag⁺ ions were considerably higher in AgCaGP/C. C. albicans was susceptible to nanocompounds produced with both reducing agents, regardless of Ag concentration and CaGP form, being Ag10%CaGP-N/C the most effective compound (19.5⁻39.0 µg Ag mL−1). While for S. mutans, the effectiveness was observed only for AgCaGP reduced by citrate, also presenting Ag10%CaGP-N the highest effectiveness (156.2⁻312.5 µg Ag mL−1). Notably, CaGP enhanced the silver antimicrobial potential in about two- and eight-fold against C. albicans and S. mutans when compared with the AgNP controls (from 7.8 to 3.9 and from 250 to 31.2 µg Ag mL−1, respectively). The synthesis that was used in this study promoted the formation of AgNP associated with CaGP, and although the use of sodium borohydride (B) resulted in a pronounced reduction of Ag⁺, the composite AgCaGP/B was less effective against the microorganisms that were tested.

Nanopartículas de prata obtidas pela síntese 'green' combinadas ou não com glicerofosfato de cálcio e tirosol: efeito antimicrobiano, anti-inflamatório e na resposta transcriptômica em patógenos orais / Silver nanoparticles obtained by the 'green' synthesis combined or not with calcium glycerophosphate and tyrosol: antimicrobial, anti-inflammatory effect and in the transcriptomic response in oral pathogens

O objetivo deste estudo foi avaliar a atividade antimicrobiana/antibiofilme e a viabilidade celular de nanopartículas de prata (NPsAg) obtidas por uma síntese 'green' associadas ou não ao ß-glicerofosfato de cálcio (GPCa) contra Streptococcus mutans e espécies de Candida (cepas de referência e isolados clínicos orais incluindo cepas resistentes ao fluconazol). O efeito destes nanocompostos em combinação com o tirosol (TIR), fluconazol (FLC), nistatina (NIT) e anfotericina B (AnB) também foi avaliado. Além disso, nós avaliamos comparativamente as alterações do transcriptoma de células de C. glabrata CBS138 após exposição às NPsAg e ao Íon prata (Ag+ ). Inicialmente, as NPsAg foram sintetizadas por meio da redução do nitrato de prata com extratos de diferentes partes (casca, folha e semente) de uma romã (Punica granatum L.) associadas ou não ao GPCa. A Concentração Inibitória Mínima (CIM) e as Concentrações Fungicida/Bactericida Mínima (CFM e CBM) dos nanocompostos (NPsAg e NPsAgGPCa) associados ou não ao TIR contra S. mutans e C. albicans foram determinadas pelo método de microdiluição. E, a ação das NPsAg combinadas com FLC, NIT e AnB também foi avaliada em condição planctônica contra isolados clínicos orais de espécies de Candida. O efeito dos nanocompostos associados ou não ao TIR sobre a viabilidade celular de fibroblastos da linhagem L929 e a produção de citocinas foi avaliado através dos ensaios de MTT e ELISA, respectivamente. O número de hifas foi quantificado em biofilmes de C. albicans formados na presença das NPsAg com ou sem soro fetal bovino (SFB). O efeito das NPsAg em inibir a formação do biofilme (em C. albicans e C. glabrata) também foi investigado através do ensaio de PrestoBlue e por meio da microscopia eletrônica de varredura. Os nanocompostos (NPsAg e NPsAg-GPCa) associados ou não ao TIR foram aplicados sobre biofilmes de S. mutans e de espécies de Candida (12 e 24 h) e, após 24 h de contato, sua atividade antibiofilme foi determinada por meio da enumeração das unidades formadoras de colônias (UFCs) e ensaio de PrestoBlue. Além disso, uma análise transcriptômica foi realizada utilizando microchips de DNA ('microarrays') a fim de avaliar quais genes estão mais ou menos expressos como resposta às NPsAg (no estado planctônico e formando biofilmes) e ao Íon Ag+ (formando biofilmes). As soluções antimicrobianas sintetizadas neste estudo (NPsAg e NPsAgGPCa) apresentaram atividade antimicrobiana contra os microrganismos testados. Além disso, menores valores de CIM foram obtidos quando estes nanocompostos foram associados ao TIR, FLC, NIT e AnB apresentando um efeito sinérgico. NPsAg e NPsAgGPCa não foram tóxicas às células L929, aumentaram a produção de fator de crescimento celular e não promoveram alterações significativas na liberação de Interleucina-6. Os nacompostos associados ao TIR não apresentaram efeito citotóxico sobre culturas de L929, exceto para a maior concentração (NPsAg ­ 39,05 µg/mL + TIR ­ 1,25 mM). A presença das NPsAg reduziu drasticamente o número de hifas em biofilmes de C. albicans na presença ou na ausência de SFB. A quantidade de biofilme das espécies de Candida formado na presença das NPsAg reduziu para mais de 50%. Após 24 h de tratamento com os nanocompostos (NPsAg e NPsAg-GPCa) em biofilmes de S. mutans, houve uma redução significativa no número de UFCs sendo similar à clorexidina. Uma redução na viabilidade de biofilmes de C. glabrata também foi observada após exposição às NPsAg (24 h). Entretanto, os nanocompostos (NPsAg e NPsAg-GPCa) associados ou não ao TIR não foram efetivos contra biofilmes de C. albicans. Após exposição às NPsAg e ao Íon Ag+ , alterações no transcriptoma de células de C. glabrata CBS138 foram observadas: no estado planctônico, houve uma superexpressão dos genes responsáveis pela biossíntese de metionina e de lisina e uma subexpressão dos genes relacionados com o transporte transmembrana; e, as células expostas às NPsAg responderam de forma distinta em relação ao Íon Ag+ , indicando que as NPsAg podem apresentar um mecanismo de ação diferente. Estes achados podem auxiliar nas decisões terapêuticas com formulações contendo NPsAg ou NPsAg-GPCa associadas ou não a diferentes drogas em pacientes com cárie dentária e candidíase oral(AU)

The aim of this study was to evaluate the antimicrobial/antibiofilm activities and the cell viability of silver nanoparticles (AgNPs) obtained by a 'green' synthesis associated or not to ß-calcium glycerophosphate (CaGP) against Streptococcus mutans and Candida species (reference strains and oral clinical isolates including azole-resistant strains). The effect of these nanocompounds in combination with tyrosol (TYR), fluconazole (FLC), nystatin (NYT) and amphotericin B (AmB) also was evaluated. Furthermore, we evaluated, comparatively, the transcriptome alterations of cells of C. glabrata CBS138 after exposition to AgNPs and to silver ion (Ag+ ). Initially, AgNPs were synthesized by reducing silver nitrate with extracts of different parts (peel, leaves and seeds) of a pomegranate (Punica granatum L.) associated or not to CaGP. Minimum Inhibitory Concentration (MIC) and Minimum Fungicidal/Bactericidal Concentrations (MFC and MBC) of the nanocomposites (AgNPs and AgNPs-CaGP) associated or not to TYR against S. mutans and C. albicans were determined by the microdilution method. And, the action of the AgNPs combined with FLC, NYT and AmB also was evaluated in planktonic condition against oral clinical isolates of Candida species. The effect of the nanocomposites associated or not to TYR on cell viability of fibroblasts (L929) and cytokines production was evaluated through MTT and ELISA assays, respectively. The number of cells undergoing filamentation was quantified in C. albicans biofilms formed in the presence of AgNPs with or without fetal bovine serum (FBS). The effect of AgNPs in inhibit the formation of biofilm (in C. albicans and C. glabrata) also was investigated through PrestoBlue assay and scanning electron microscope. Biofilms of S. mutans and Candida species (12 and 24 h) were treated with nanocomposites (AgNPs and AgNPsCaGP) associated or not to TYR for 24 h and, then, the viability of these biofilms was determined through PrestoBlue assay and colony forming units (CFUs). Moreover, a transcriptome analysis was performed using microarrays to determine which genes are up- or down-regulated as response to AgNPs (in the planktonic state and forming biofilms) and to Ag+ ion (forming biofilms). Antimicrobial solutions synthesized in this study (AgNPs and AgNPs-CaGP) presented antimicrobial activity against tested microorganisms. Furthermore, lower values of MIC were obtained when these nanocompounds were associated to TYR, FLC, NYT and AmB showing a synergistic effect. AgNPs and AgNPs-CaGP were not toxic to L929 cells, increased the stem cell factor production and did not promote significant alterations in the Interleukine-6 release. The nanocomposites associated to TYR did not present cytotoxic effect on L929 cultures, except for the higher concentration (AgNPs ­ 39.05 µg/mL + TYR ­ 1.25 mM). The incubation in the presence of the AgNPs drastically reduced the number of cells exhibiting hyphae, this effect being observed either in the presence or absence of FBS. The amount of biofilm formed by Candida species in the presence of the AgNPs was reduced by more than 50% the one formed in the absence of the nanoparticles, this reduction being further increased to more than 90% when the concentration of the AgNPs was increased. After 24 h of treatment with the nanocompounds (AgNPs and AgNPs-CaGP) in S. mutans biofilms, there was a significant reduction in the number of CFUs being similar to chlorhexidine. A reduction in the viability of C. glabrata biofilms also was observed after exposition to AgNPs (24 h). However, the nanocomposites (AgNPs and AgNPs-CaGP) associated or not to TYR were not effective against C. albicans biofilms. After exposition to AgNPs and to Ag+ ion, alterations in the transcriptome of cells of C. glabrata CBS138 were observed: in the planktonic state, genes responsible by the methionine and lysine biosynthesis are up-regulated, and genes related with the transmembrane transport are down-regulated. And, finally, the cells exposed to AgNPs responded differently in relation to the Ag+ ion, indicating that the AgNPs might present a different mechanism of action. All these results may help guide therapeutic decisions with formulation containing AgNPs or AgNPs-CaGP associated or not with different compounds in patients with dental caries and oral candidiasis(AU)

Fluoride toothpaste supplemented with sodium hexametaphosphate reduces enamel demineralization in vitro.

OBJECTIVE: The aim of this study was to evaluate the effect of fluoride dentifrices combined with sodium hexametaphosphate (HMP) on enamel demineralization in vitro. MATERIAL AND METHODS: Enamel bovine blocks were selected by initial surface hardness (SHi) and then divided into five experimental groups (n = 12): placebo (without fluoride and without HMP); 1100 ppm F (1100F); and 1100F associated with HMP at 0.5 % (1100HMP0.5%), 1 % (1100HMP1%), and 2 % of HMP (1100HMP2%). Blocks were submitted to five pH cycles (demineralizing/remineralizing solutions) at 37 °C. After pH cycling, final surface hardness (SHf), percentage of surface hardness loss (%SH), integrated differential hardness (ΔIH), integrated loss of subsurface hardness (ΔKHN), and enamel firmly bound fluoride (F) were determined. Data were submitted to one-way ANOVA, followed by Student-Newman-Keuls test (p < 0.05). RESULTS: Significant differences were observed among all groups regarding %SH and ΔKHN. 1100HMP1% promoted the lowest mineral loss among all groups (p < 0.001), and led to significantly lower demineralization in the deeper regions of the subsurface lesion when compared with the other HMP-containing toothpastes (p < 0.001). Significantly higher mineral loss was observed for 1100HMP2% when compared to the other fluoridated dentifrices, mainly in the outer part of the lesion (p < 0.001). Enamel F uptake was similar for 1100F and 1100HMP1% but significantly reduced for other HMP concentrations. CONCLUSION: The supplementation of a 1100-ppm F dentifrice with 1 % HMP promoted a higher inhibitory effect against enamel demineralization when compared to a dentifrice containing the same amount of fluoride in vitro. CLINICAL RELEVANCE: This dentifrice could potentially be indicated to patients at high risk of caries.

Effect of Trimetaphosphate and Fluoride Association on Hydroxyapatite Dissolution and Precipitation In Vitro

The present study analyzed the action of sodium trimetaphosphate (TMP) and/or fluoride on hydroxyapatite. Hydroxyapatite powder was suspended in different solutions: deionized water, 500 µg F/mL, 1,100 µg F/mL, 1%TMP, 3%TMP, 500 µg F/mL plus 1%TMP and 500 µg F/mL plus 3%TMP. The pH value of the solutions was reduced to 4.0 and after 30 min, raised to 7.0 (three times). After pH-cycling, the samples were analyzed by X-ray diffraction and infrared spectroscopy. The concentrations of calcium fluoride, fluoride, calcium and phosphorus were also determined. Adding 1% or 3% TMP to the solution containing 500 µg F/mL produced a higher quantity of calcium fluoride compared to samples prepared in a 1,100 µg F/mL solution. Regarding the calcium concentration, samples prepared in solutions of 1,100 µg F/mL and 500 µg F/mL plus TMP were statistically similar and showed higher values. Using solutions of 1,100 µg F/mL and 500 µg F/mL plus TMP resulted in a calcium/phosphorus ratio close to that of hydroxyapatite. It is concluded that the association of TMP and fluoride favored the precipitation of a more stable hydroxyapatite.

O presente estudo avaliou a ação do trimetafosfato de sódio (TMP) e/ou fluoreto sobre a hidroxiapatita. Pó de hidroxiapatita foi suspenso em diferentes soluções: água deionizada, 500 µg F/mL, 1100 µg F/mL, 1%TMP, 3%TMP, 500 µg F/mL adicionado a 1%TMP e 500 µg F/mL associado a 3%TMP. O pH das soluções foi reduzido para 4,0 e depois de 30 min, elevado para 7,0 (três vezes). Depois do processo de ciclagem de pH, as amostras foram analisadas por difração de raios-X e espectroscopia por infravermelho. As concentrações de fluoreto de cálcio, fluoreto, cálcio e fósforo também foram determinadas. A adição de 1% ou 3% TMP na solução contendo 500 µg F/mL produziu uma maior quantidade de fluoreto de cálcio comparado às amostras tratadas com uma solução de 1100 µg F/mL. A respeito da concentração de cálcio, amostras tratadas com soluções de 1100 µg F/mL e 500 µg F/mL adicionado ao TMP foram estatisticamente similares e mostraram maiores valores. Soluções de 1100 µg F/mL e 500 µg F/mL adicionado ao TMP resultaram em uma proporção molar Ca/P mais próxima à da hidroxiapatita. Conclui-se que a associação de TMP e F favoreceu a precipitação de uma hidroxiapatita mais estável.

Effect of trimetaphosphate and fluoride association on hydroxyapatite dissolution and precipitation in vitro.

The present study analyzed the action of sodium trimetaphosphate (TMP) and/or fluoride on hydroxyapatite. Hydroxyapatite powder was suspended in different solutions: deionized water, 500 µg F/mL, 1,100 µg F/mL, 1%TMP, 3%TMP, 500 µg F/mL plus 1%TMP and 500 µg F/mL plus 3%TMP. The pH value of the solutions was reduced to 4.0 and after 30 min, raised to 7.0 (three times). After pH-cycling, the samples were analyzed by X-ray diffraction and infrared spectroscopy. The concentrations of calcium fluoride, fluoride, calcium and phosphorus were also determined. Adding 1% or 3% TMP to the solution containing 500 µg F/mL produced a higher quantity of calcium fluoride compared to samples prepared in a 1,100 µg F/mL solution. Regarding the calcium concentration, samples prepared in solutions of 1,100 µg F/mL and 500 µg F/mL plus TMP were statistically similar and showed higher values. Using solutions of 1,100 µg F/mL and 500 µg F/mL plus TMP resulted in a calcium/phosphorus ratio close to that of hydroxyapatite. It is concluded that the association of TMP and fluoride favored the precipitation of a more stable hydroxyapatite.

Efeito do trimetafosfato de sódio e do fluoreto sobre a solubilidade da hidroxiapatita: estudo in vitro / Effect of sodium trimetaphosphate and fluoride on hydroxyapatite solubility: an in vitro study

O objetivo deste estudo foi avaliar o efeito de diferentes concentrações de trimetafosfato de sódio (TMP) associado ao fluoreto (F) sobre a solubilidade da hidroxiapatita (HA), bem como as propriedades físicas e químicas da HA depois de um ciclo de pH. O pó de HA sintética (1,0 g; n= 6) foi misturado com soluções de TMP nas concentrações de 0; 0,1; 0,2; 0,4; 0,6; 0,8; 1,0; 2,0; 4,0; 6,0; 8,0 e 10%, associadas ao F nas concentrações de 0, 100, 250 e 500 ppm F, durante 2 min. A suspensão foi filtrada e o precitado foi seco por 24 horas a 37°C e moído. Depois do tratamento, as amostras de HA tratada foram submetidas a um ciclo de pH. As amostras de HA ciclada foram caracterizadas por energia dispersiva de raios-X (EDX), espectroscopia de infravermelho (FTIR) e difração de raios-X (DRX). Depois, as concentrações de F solúvel em base e em ácido, cálcio (Ca) e fósforo (P) foram determinadas na HA ciclada, assim como as concentrações de P e F no sobrenadante. Os dados de F, Ca, P e proporção molar Ca/P na HA, e F e P no sobrenadante foram submetidas à Análise de Variância e teste de Student-Newman-Keuls (p<0,05). Houve uma redução do tamanho dos cristais de HA com o aumento da concentração de TMP. A HA tratada com 500 ppm F e TMP a 1% produziu uma HA com cristalitos de tamanho maiores. O FTIR mostrou uma redução na intensidade das bandas de fosfatos e carbonato quando comparado à HA sintetizada. As maiores proporções Ca/P foram observadas na HA tratada com TMP entre 0,4% e 1% quando combinado com 250 ppm F, e entre 0,4% e 2% em associação com 500 ppm F (p<0,05). Um aumento na concentração de TMP levou a uma redução na incorporação de F solúvel em ácido na estrutura do cristal para todos os grupos, nas duas situações (depois do tratamento e depois do ciclo de pH) (p<0,05). Adicionalmente, um aumento na concentração de TMP também proporcionou uma maior adsorção de TMP à HA para as soluções de 0 e 100 ppm F, mas menor para as soluções de 250 e 500 ppm F (p<0,05)...

The aim of this study was to evaluate the effect of different concentrations of sodium trimetaphosphate (TMP) associated with fluoride (F) on hydroxyapatite (HA) solubility, as well as in chemical and physical properties of HA after a pH-cycle. Synthetic HA powder (1.0 g; n= 6) was mixed with solutions of TMP at 0, 0.1, 0.2, 0.4, 0.6, 0.8, 1.0, 2.0, 4.0, 6.0, 8.0 and 10%, associated with fluoride at 0, 100, 250 and 500 ppm F, during 2 min. The suspension was filtered and the precipitates were dried for 24 h at 37°C and grinded. After the treatment, HA samples were submitted to a pH-cycle. Post-cycled HA samples were analyzed by energy-dispersive X-ray spectroscopy (EDX), infrared spectroscopy (FTIR) and X-ray diffraction (XRD). Afterwards, the concentration of alkali and acid soluble F, Ca and P were determined in post-cycled HA, as well as P and F in the supernatant. The data of F, Ca, P, Ca/P ratio of HA, and F and P in the supernatant were submitted to ANOVA and Student-Newman-Keuls’ test (p<0.05). A reduction of the size of the HA crystallites was seen with increasing TMP concentrations in the solutions. HA treated with 500 ppm F and 1% TMP produced an HA with crystallites of larger size. The FTIR showed a reduction in the bands corresponding to phosphates and to carbonate for all groups when compared to the HA synthetized. The highest Ca/P ratios were observed for HA treated with TMP concentrations between 0.4% and 0.8% when combined with 250 ppm F, and between 0.4% and 2% in association with 500 ppm F (p<0.05). The increase of TMP led a reduced acid-soluble F incorporation in HA for all groups, both immediately after treatment and after the pH cycle (p<0.05). Additionally, the increase in TMP concentrations led to higher P adsorption to HA for the 0 and 100 ppm F solutions, but lower for the 250 and 500 ppm F solutions (p<0,05). The results indicate that the mechanism of action of TMP seems to be related with the TMP adsorption to crystal structural...

Efeito do trimetafosfato de sódio e do fluoreto sobre a solubilidade da hidroxiapatita: estudo in vitro / Effect of sodium trimetaphosphate and fluoride on hydroxyapatite solubility: an in vitro study

O objetivo deste estudo foi avaliar o efeito de diferentes concentrações de trimetafosfato de sódio (TMP) associado ao fluoreto (F) sobre a solubilidade da hidroxiapatita (HA), bem como as propriedades físicas e químicas da HA depois de um ciclo de pH. O pó de HA sintética (1,0 g; n= 6) foi misturado com soluções de TMP nas concentrações de 0; 0,1; 0,2; 0,4; 0,6; 0,8; 1,0; 2,0; 4,0; 6,0; 8,0 e 10%, associadas ao F nas concentrações de 0, 100, 250 e 500 ppm F, durante 2 min. A suspensão foi filtrada e o precitado foi seco por 24 horas a 37°C e moído. Depois do tratamento, as amostras de HA tratada foram submetidas a um ciclo de pH. As amostras de HA ciclada foram caracterizadas por energia dispersiva de raios-X (EDX), espectroscopia de infravermelho (FTIR) e difração de raios-X (DRX). Depois, as concentrações de F solúvel em base e em ácido, cálcio (Ca) e fósforo (P) foram determinadas na HA ciclada, assim como as concentrações de P e F no sobrenadante. Os dados de F, Ca, P e proporção molar Ca/P na HA, e F e P no sobrenadante foram submetidas à Análise de Variância e teste de Student-Newman-Keuls (p<0,05). Houve uma redução do tamanho dos cristais de HA com o aumento da concentração de TMP. A HA tratada com 500 ppm F e TMP a 1% produziu uma HA com cristalitos de tamanho maiores. O FTIR mostrou uma redução na intensidade das bandas de fosfatos e carbonato quando comparado à HA sintetizada. As maiores proporções Ca/P foram observadas na HA tratada com TMP entre 0,4% e 1% quando combinado com 250 ppm F, e entre 0,4% e 2% em associação com 500 ppm F (p<0,05). Um aumento na concentração de TMP levou a uma redução na incorporação de F solúvel em ácido na estrutura do cristal para todos os grupos, nas duas situações (depois do tratamento e depois do ciclo de pH) (p<0,05). Adicionalmente, um aumento na concentração de TMP também proporcionou uma maior adsorção de TMP à HA para as soluções de 0 e 100 ppm F, mas menor para as soluções de 250 e 500 ppm F (p<0,05)...

The aim of this study was to evaluate the effect of different concentrations of sodium trimetaphosphate (TMP) associated with fluoride (F) on hydroxyapatite (HA) solubility, as well as in chemical and physical properties of HA after a pH-cycle. Synthetic HA powder (1.0 g; n= 6) was mixed with solutions of TMP at 0, 0.1, 0.2, 0.4, 0.6, 0.8, 1.0, 2.0, 4.0, 6.0, 8.0 and 10%, associated with fluoride at 0, 100, 250 and 500 ppm F, during 2 min. The suspension was filtered and the precipitates were dried for 24 h at 37°C and grinded. After the treatment, HA samples were submitted to a pH-cycle. Post-cycled HA samples were analyzed by energy-dispersive X-ray spectroscopy (EDX), infrared spectroscopy (FTIR) and X-ray diffraction (XRD). Afterwards, the concentration of alkali and acid soluble F, Ca and P were determined in post-cycled HA, as well as P and F in the supernatant. The data of F, Ca, P, Ca/P ratio of HA, and F and P in the supernatant were submitted to ANOVA and Student-Newman-Keuls’ test (p<0.05). A reduction of the size of the HA crystallites was seen with increasing TMP concentrations in the solutions. HA treated with 500 ppm F and 1% TMP produced an HA with crystallites of larger size. The FTIR showed a reduction in the bands corresponding to phosphates and to carbonate for all groups when compared to the HA synthetized. The highest Ca/P ratios were observed for HA treated with TMP concentrations between 0.4% and 0.8% when combined with 250 ppm F, and between 0.4% and 2% in association with 500 ppm F (p<0.05). The increase of TMP led a reduced acid-soluble F incorporation in HA for all groups, both immediately after treatment and after the pH cycle (p<0.05). Additionally, the increase in TMP concentrations led to higher P adsorption to HA for the 0 and 100 ppm F solutions, but lower for the 250 and 500 ppm F solutions (p<0,05). The results indicate that the mechanism of action of TMP seems to be related with the TMP adsorption to crystal structural of...

Effect of sodium trimetaphosphate on hydroxyapatite solubility: an in vitro study.

This study evaluated the effect of different concentrations of sodium trimetaphosphate (TMP) with and without fluoride (F) on the concentration of calcium (Ca), phosphorus (P) and F in hydroxyapatite (HA). Synthetic HA powder (0.15 g) was suspended (n=6) in solutions (75 mL) of TMP at 0%, 0.1%, 0.2%, 0.4%, 0.6%, 0.8%, 1.0%, 2.0%, 4.0%, 6.0%, 8.0% and 10% concentrations in the presence and absence of 100 ppm F and subjected to a pH-cycling process. The precipitates were filtrated, dried at 70° C for 24 h and ground onto a fine powder. The concentrations of F (KOH (CaF2) and HCl (FA) soluble), Ca (Arsenazo III), and P (molybdate method) in HA were determined. The Ca P, and Ca/P ratio data were subjected to Tukey's test and the F data were subjected to Student-Newman-Keuls test (p<0.05). The addition of TMP to the samples reduced F deposition to 98% (p<0.001). The groups containing 100 ppm F and 0.4% or 0.6% TMP exhibited a higher Ca concentration than the group containing only 100 ppm F (p<0.05). Furthermore, the HA treated with 0.2% and 0.4% TMP and 100 ppm F showed a higher Ca/P ratio than the other groups (p<0.001). In conclusion, TMP at 0.2%, 0.4% and 0.6% concentrations combined with F seemed to be able to precipitate HA with low solubility. However, especially at high concentrations, TMP interferes with F deposition on HA.

Effect of Sodium Trimetaphosphate on Hydroxyapatite Solubility: An In Vitro Study

This study evaluated the effect of different concentrations of sodium trimetaphosphate (TMP) with and without fluoride (F) on the concentration of calcium (Ca), phosphorus (P) and F in hydroxyapatite (HA). Synthetic HA powder (0.15 g) was suspended (n=6) in solutions (75 mL) of TMP at 0%, 0.1%, 0.2%, 0.4%, 0.6%, 0.8%, 1.0%, 2.0%, 4.0%, 6.0%, 8.0% and 10% concentrations in the presence and absence of 100 ppm F and subjected to a pH-cycling process. The precipitates were filtrated, dried at 70° C for 24 h and ground onto a fine powder. The concentrations of F (KOH (CaF2) and HCl (FA) soluble), Ca (Arsenazo III), and P (molybdate method) in HA were determined. The Ca P, and Ca/P ratio data were subjected to Tukey's test and the F data were subjected to Student-Newman-Keuls test (p<0.05). The addition of TMP to the samples reduced F deposition to 98% (p<0.001). The groups containing 100 ppm F and 0.4% or 0.6% TMP exhibited a higher Ca concentration than the group containing only 100 ppm F (p<0.05). Furthermore, the HA treated with 0.2% and 0.4% TMP and 100 ppm F showed a higher Ca/P ratio than the other groups (p<0.001). In conclusion, TMP at 0.2%, 0.4% and 0.6% concentrations combined with F seemed to be able to precipitate HA with low solubility. However, especially at high concentrations, TMP interferes with F deposition on HA.

Este estudo avaliou o efeito de diferentes concentrações de trimetafosfato de sódio (TMP) com e sem fluoreto (F) nas concentrações de cálcio (Ca), fósforo (P) e F na hidroxiapatita (HA). Pó de HA sintético (0,15 g) foi suspenso (n=6) em soluções (75 mL) de TMP com concentrações de 0%, 0,1%, 0,2%, 0,4%, 0,6%, 0,8%, 1,0%, 2,0%, 4,0%, 6,0%, 8,0% e 10% na presença ou na ausência de 100 ppm F e foram submetidas ao processo de ciclagem de pH. O precipitado foi filtrado seco a 70°C por 24 h e triturado em um pó fino. As concentrações de F (solúvel em KOH: CaF2, e em HCl: FA), Ca (Arsenazo III) e P (método do molibdato) foram determinadas na HA. Os dados de Ca, P e de proporção Ca/P foram submetidos ao teste de Tukey e os dados de F ao teste Student-Newman-Keuls (p<0,05). A adição de TMP reduziu a deposição de F em 98% (p<0,001). Os grupos contendo 100 ppm F e TMP 0,4% e 0,6% apresentaram maiores concentrações de Ca do que o grupo contendo somente 100 ppm F (p<0,05). Além disso, a HA tratada com 0,2% e 0,4% de TMP e 100 ppm F apresentou maiores proporções Ca/P em relação aos demais grupos (p<0,001). Em conclusão, TMP nas concentrações de 0,2%, 0,4% e 0,6% quando associado ao F é capaz de precipitar uma HA com menor solubilidade. Entretanto, especialmente em altas concentrações, TMP interfere com a deposição de F na HA.