Enhancement of aqueous solubility and antibiofilm activity of 4-allylpyrocatechol by polymeric micelles.

4-Allylpyrocatechol (APC), a major active compound of Piper betle, possesses strong antimicrobial activity. However, the water-insoluble property of APC limits its clinical and pharmaceutical use. To solve this problem, APC loaded polymeric micelles (PMAC) was fabricated using the thin-film hydration method. Nanoparticles of PMAC were characterized using a photon correlation spectrophotometer and transmission electron microscope (TEM). Antibiofilm activity of PMAC was investigated using crystal violet assay and confocal laser scanning microscopy (CLSM). Cytotoxic effects of PMAC on normal cells were investigated using MTT assay. The results demonstrate that a ratio of APC to the polymer plays an important role in the physicochemical characteristics of PMAC. The most suitable PMAC formulation having a small particle size (38.8 ± 1.4 nm), narrow size distribution (0.28 ± 0.10), a high negative zeta potential (- 16.43 ± 0.55 mV), and high entrapment efficiency (86.33 ± 14.27%) can be obtained from the ratio 1:4. The water solubility of this PMAC is significantly improved, approximately 1,000-fold higher than the unentrapped APC. TEM images demonstrate that PMAC is spherical in shape. The inhibitory effects of PMAC (1.5 mg APC/mL) against Streptococcus intermedius and Streptococcus mutans biofilms are significantly stronger than chlorhexidine (0.06 mg/mL). Images from CLSM demonstrate the destruction and thickness reduction of the pathogenic biofilms after contacting with PMAC. The MTT assay confirms that PMAC at this concentration is non-toxic to normal cells. These results obviously indicate that PMAC is a promising natural and harmless antimicrobial agent suitable for use in the oral cavity for inhibition of pathogenic bacterial biofilms.

Anesthetic efficacies of intrapapillary injection in comparison to inferior alveolar nerve block for mandibular premolar extraction: a randomized clinical trial.

OBJECTIVE: Intrapapillary injection (IPI) has been suggested to improve pulpal anesthesia of mandibular teeth and to avoid complications from inferior alveolar nerve block (IANB). This study aimed to determine and compare clinical efficacies and prostaglandin E2 (PGE2) levels between IPI and IANB. MATERIALS AND METHODS: IANB was randomly selected for mandibular premolar anesthesia on one side of 40 patients, whereas IPI was locally administered to the contralateral premolar. Pulpal anesthesia, pain during injection and extraction, patients' satisfaction, and complications were assessed from 30 patients. Gingival crevicular fluid from ten patients was collected for PGE2 quantification by ELISA. RESULTS: Of 30 patients, 18 preferred IPI after injection due to significantly faster mean onset of pulpal anesthesia (p < 0.001) and lower mean score of injection pain (p = 0.017) than IANB, but 21 preferred IANB instead after extraction due to less postoperative pain, consistent with the significantly lower median PGE2 level on the IANB side than that on the IPI at 30 min (p = 0.047). However, there was no difference in the mean satisfaction score between the two techniques. Ulcerated epithelium and sloughing tissues were found at the IPI site in some patients with complete healing within 2 weeks. CONCLUSIONS: The anesthetic efficacies of IPI for mandibular premolar extraction are comparable to those of IANB. However, postoperative pain and local complications at the IPI site should be considered. CLINICAL RELEVANCE: IPI may be used for dental procedures that require only a short anesthetic duration to avoid failure of pulpal anesthesia, complications, and discomfort from IANB.

Positive correlations between hCAP18/LL-37 and chondroitin sulphate levels in chronic periodontitis.

AIM: To measure the levels of hCAP18/LL-37 in gingival crevicular fluid from patients with periodontal diseases compared with healthy controls and to determine the correlation between hCAP18/LL-37 and chondroitin sulphate (CS) levels in patients with periodontitis. MATERIAL AND METHODS: Gingival crevicular fluid samples from 51 patients and 25 healthy volunteers were analysed for the hCAP18/LL-37 levels by immunoblotting and were determined for the CS levels by the competitive enzyme-linked immunosorbent assay. RESULTS: Tris buffer pH 9.85 was selected to recover hCAP18/LL-37 from Periopaper strips, in which the percentages of recovery were around 70%. The median levels of hCAP18/LL-37 in the aggressive and the chronic periodontitis (CP) groups were significantly greater than those in the gingivitis and the healthy groups (p < 0.05). Significant correlations between the unprocessed 18-kDa fragment and CS levels (r = 0.650; p < 0.001) and between the mature 4.6-kDa fragment and CS levels (r = 0.502; p < 0.001) were observed only in the CP group. CONCLUSION: The significant correlations between the hCAP18/LL-37 and the CS levels were found in CP, but not in aggressive periodontitis. The presence versus absence of such correlations may be clinically applicable to help clinicians distinguish between two distinct types of periodontitis.

Favorable interleukin-8 induction in human gingival epithelial cells by the antimicrobial peptide LL-37.

BACKGROUND: LL-37, the only member of the antimicrobial peptide cathelicidin family in humans, exerts a variety of biological activities, especially immunomodulation through either direct chemotactic activity or up-regulation of several cytokines and chemokines in various cell types. In this study, we aimed to determine the immunoregulatory effect of LL-37 on Th1/Th2 cytokine expression and production in human gingival epithelial cells (HGECs). METHODS: Cultured HGECs were treated with different concentrations of LL-37 for different numbers of times. The cytotoxicity of LL-37 was determined by an MTT assay. Total RNA was isolated for RT-PCR and real-time PCR analyses of cytokine expression. Cell-free culture supernatants were assayed for Th1/Th2 cytokine levels by a cytokine bead array. RESULTS: Out of eleven Th1/Th2 cytokines tested, treatment of HGECs with non-toxic doses of LL-37 (2-6 µM) significantly raised only IL-8 levels in the cell-free culture supernatants, when compared to control untreated cells (P <0.05). Consistent with the elevated IL-8 levels, IL-8 mRNA expression was remarkably and significantly induced by LL-37 treatment (P < 0.05), when compared to the modest mRNA induction of other three cytokines, including IL-1ß, IL-6, and TNF-α. The time-course study demonstrated a cumulative IL-8 mRNA induction by LL-37 treatment within a 24-hour interval. CONCLUSIONS: These findings indicate that LL-37 favorably induces IL-8 expression and secretion in HGECs, suggesting both direct and indirect involvement of LL-37 in neutrophil recruitment into an inflammatory site within diseased periodontal tissues.

Comparisons between two biochemical markers in evaluating periodontal disease severity: a cross-sectional study.

BACKGROUND: The purpose of this study was to compare two biochemical markers, which have been previously used to determine the degrees of alveolar bone destruction, in evaluating periodontal disease severity. METHODS: The WF6 epitope of chondroitin sulfate (CS) and the alkaline phosphatase (ALP) levels were determined in gingival crevicular fluid (GCF) samples collected from patients with various degrees of disease severity, including ten patients with gingivitis (50 gingivitis sites) and 33 patients with chronic periodontitis (including gingivitis, slight, moderate, and severe periodontitis sites; n = 50 each), as well as from ten healthy volunteers (50 healthy sites) by Periopaper strips. The levels of CS and ALP were measured by an ELISA and a fluorometric assay, respectively. RESULTS: The results demonstrated low levels of CS and ALP in non-destructive and slightly destructive periodontitis sites, whereas significantly high levels of these two biomolecules were shown in moderately and severely destructive sites (p < 0.05). Although a significant difference in CS levels was found between moderate and severe periodontitis sites, no difference in ALP levels was found. Stronger correlations were found between CS levels and periodontal parameters, including probing depth, loss of clinical attachment levels, gingival index and plaque index, than between ALP levels and these parameters. CONCLUSIONS: It is suggested that the CS level is a better diagnostic marker than the ALP level for evaluating distinct severity of chronic periodontitis.

Development of Controlled-Release Carbamide Peroxide Loaded Nanoemulgel for Tooth Bleaching: In Vitro and Ex Vivo Studies.

Burst release of carbamide peroxide (CP) from traditional hydrogels causes severe inflammation to periodontal tissues. The present study explores the development of a novel CP nanoemulgel (CP-NG), an oil-in-water nanoemulsion-based gel in which CP was loaded with a view to controlling CP release. CP solid dispersions were prepared, using white soft paraffin or polyvinylpyrrolidone-white soft paraffin mixture as a carrier, prior to formulating nanoemulsions. It was found that carrier type and the ratio of CP to carrier affected drug crystallinity. Nanoemulsions formulated from the optimized CP solid dispersions were used to prepare CP-NG. It was found that the ratio of drug to carrier in CP solid dispersions affected the particle size and zeta potential of the nanoemulsions as well as drug release behavior and tooth bleaching efficacy of CP-NG. Drug release from CP-NG followed a first-order kinetic reaction and the release mechanism was an anomalous transport. Drug release rate decreased with an increase in solid dispersion carriers. CP-NG obtained from the solid dispersion with a 1:1 ratio of CP to the polymer mixture is suitable for sustaining drug release with high tooth bleaching efficacy and without reduction of enamel microhardness. The developed CP-NG is a promising potential tooth bleaching formulation.

Denture-Soaking Solution Containing Piper betle Extract-Loaded Polymeric Micelles; Inhibition of Candida albicans, Clinical Study, and Effects on Denture Base Resin.

Candida albicans is a common overgrowth in people wearing dentures. Long-term use of antifungal chemicals carries a risk of toxic side effects. This study focused on the edible Piper betle extract because of its safety. The broth dilution method was applied for antifungal determination of the ethyl acetate fractionated extract (fEA) and fEA-loaded polymeric micelles (PMF). The PMF was prepared by thin-film hydration using poloxamer 407 as a polymer base. The results found that the weight ratio of fEA to polymer is the main factor to obtain PMF system as a clear solution, nanoparticle sizes, narrow size distribution, negative zeta potential, and high entrapment efficiency. The activity of PMF against C. albicans is significantly higher than fEA alone, with a minimum fungicidal concentration of 1.5 mg/mL. PMF from 1:3 ratio of fEA to polymer is used to develop a denture-soaking solution contained 1.5 mg fEA/mL (PMFS). A clinical study on dentures of 15 volunteers demonstrated an 86.1 ± 9.2% reduction of C. albicans after soaking the dentures in PMFS daily for 14 days. Interestingly, PMFS did not change the hardness and roughness of the denture base resins. The developed PMFS may serve as a potential natural denture-soaking solution against candidiasis in denture wearers.

Development of Self-Nanoemulsifying Drug Delivery Systems Containing 4-Allylpyrocatechol for Treatment of Oral Infections Caused by Candida albicans.

Clinical use of 4-Allylpyrocatechol (APC), a potential antifungal agent from Piper betle, is limited because of its low water solubility. The current study explores the development of the self-nanoemulsifying drug delivery system (SNEDDS) containing APC (APC-SNEDDS) to enhance APC solubility. Results demonstrated that excipient type and concentration played an important role in the solubility of APC in the obtained SNEEDS. SNEDDS, comprising 20% Miglyol 812N, 30% Maisine 35-1, 40% Kolliphor RH40, and 10% absolute ethanol, provided the highest loading capacity and significantly increased water solubility of APC. Oil-in-water nanoemulsions (NE) with droplet sizes of less than 40 nm and a narrow size distribution were obtained after dispersing this APC-SNEDDS in water. The droplets had a negative zeta potential between -10 and -20 mV. The release kinetics of APC from APC-SNEDDS followed the Higuchi model. The NE containing 1.6 mg APC/mL had effective activity against Candida albicans with dose-dependent killing kinetics and was nontoxic to normal cells. The antifungal potential was similar to that of 1 mg nystatin/mL. These findings suggest that APC-SNEDDS are a useful system to enhance the apparent water solubility of APC and are a promising system for clinical treatment of oral infection caused by C. albicans.

Comparative inhibitory effects of 4-allylpyrocatechol isolated from Piper betle on Streptococcus intermedius, Streptococcus mutans, and Candida albicans.

Streptococcus intermedius, Streptococcus mutans, and Candida albicans are harmful oral pathogens and prone to resist chemical antimicrobial agents. Active ingredients from plants are of increasing interest as an alternative. This study aims to compare antimicrobial effects of 4-allylpyrocatechol (APC) extracted from Piper betle on these oral pathogens. Minimum concentration of APC against the tested pathogens was determined using a broth microdilution assay. Killing kinetic study of APC was carried out within 24 h. Morphology of the pathogenic cells was assessed using scanning electron microscopy (SEM). Anti-biofilm was investigated using crystal violet assay and confocal laser scanning microscopy (CLSM). The results showed that the mechanism of inhibition of APC was bactericidal and fungicidal effects. APC at minimum concentration of 400 µg/mL could completely kill Streptococcus and Candida spp., however, the killing rate on S. intermedius and C. albicans was significantly faster than on S. mutans. APC inhibited biofilm formation of C. albicans more efficiently than that of the bacterial cells. Cell morphology from SEM indicated that APC caused bacterial cell membrane destruction and inhibited fungal budding or tubing formation. CLSM images confirmed the killing potential of APC and suggested that bacterial dead cells could be easier washed out than the fungal dead cells. It is concluded that APC potentially inhibits growth and biofilms of oral Streptococcus and Candida spp. in different mechanism of action and killing rate. APC can be considered as a promising agent for preventing and treating dental disorders caused by S. intermedius, S. mutans, and C. albicans.

Enhancing Stability and Tooth Bleaching Activity of Carbamide Peroxide by Electrospun Nanofibrous Film.

Carbamide peroxide (CP) possesses a strong tooth bleaching activity, however, its clinical application is limited because of its instability in aqueous formulations. This study explores the improvement of CP stability and bleaching activity by loading CP in electrospun nanofibrous film (ENF). Polyvinylalcohol, polyvinylpyrrolidone, and silica were used as components for core-based nanofibers of ENF. Electrospinning feed aqueous solutions (EFASs) were developed for preparing CP loaded ENF (CP-ENF). Stability of CP in EFASs is significantly higher than in pure water. The highest stability of CP is found in PPS-CP3, composed of 0.5% CP, 5.5% polyvinylalcohol, 3% polyvinylpyrrolidone, and 1% silica. The results from X-ray diffraction indicate that CP is dispersed as a non-crystalline form in CP-ENFs. CP and the compositions of EFASs play a major role on characteristics and bleaching efficiency of CP-ENFs. Drug release of CP-ENFs is the first order kinetics. CP-ENF obtained from PPS-CP3 shows the highest drug entrapment efficiency, high adhesion, and suitable sustained release. Drug release mechanism is along with anomalous transport according to Korsmeyer-Peppas model. In an ex vivo study using human teeth, it shows the highest bleaching efficiency among the others. Therefore, CP-ENF obtained from PPS-CP3 is a promising ENF for clinical use.

Antifungal activity of polymeric micelles of silver nanoparticles prepared from Psidium guajava aqueous extract.

In the present study, silver nanoparticles (AgNPs) were synthesized by green synthesis using Psidium guajava aqueous extract (PE) as a reducing agent and silver nitrate (AgNO3) as a precursor. The obtained AgNPs showed maximum absorbance at 455 nm. The results from energy-dispersive X-ray spectroscopy demonstrate Ag signal at 88.33% weight. The particle image under scanning electron microscopy is spherical shape. The average size of the freshly prepared AgNPs is 96 ± 4 nm but is dramatically increases during storage due to particle aggregation. Coating AgNPs with polymeric micelles of poloxamer 407 (F127) at the suitable ratio can decrease the size of the freshly prepared AgNPs to 70.4 ± 0.8 nm and significantly prevent AgNPs from aggregation. The obtained coated AgNPs showed high effective on inhibition of Candida albicans. Isotonic solutions of 0.9% NaCl and phosphate buffer solution pH 7.4 can cause some extend of aggregation and increase the particle size of the coated AgNPs but the increased size is in the colloidal range that no precipitation occurs during 90 days at room temperature. From our results, it is suggested that the 1:1 ratio of AgNPs/F127 is the most suitable ratio to obtain the AgNPs loaded polymeric micelles with high stability, small particle size, and high inhibitory activity against C. albicans. These AgNPs are the promising antifungal nanomaterials for further study in animal model.

Silver nanoparticles fabricated by reducing property of cellulose derivatives.

The aim of this study was to synthesize silver nanoparticles (AgNPs) by using cellulose derivatives as a reducing agent. Methyl cellulose (MC), hydroxy ethylcellulose (HEC), and hydroxypropyl methylcellulose (HPMC) were compared for their reducing property. HPMC presented the highest reducing power, with equilibrium concentration (EC) of 84.6 ± 4.5 µmol Fe2+/g, followed by MC and HEC, with the EC of 62.3 ± 1.4, and 38.1 ± 3.2 µmol Fe2+/g, respectively. Using these cellulose derivatives as a reducing agent and silver nitrate as a precursor in fabrication of silver nanoparticles (AgNPs), three cellulose-AgNPs, HEC-AgNPs, MC-AgNPs, and HPMC-AgNPs, were obtained. The cellulose-AgNPs showed different maximum absorptions confirming AgNPs spectra at 415, 425, and 418 nm, respectively. Reaction parameters such as pH, temperature, and period of reaction affected intensity of the maximum absorptions and size of AgNPs. Using 0.3% cellulose solution at pH 9 and reaction at 70°C for 90 min, the particle size of MC-AgNPs, HEC-AgNPs, and HPMC-AgNPs was 97.7 ± 2.4, 165.6 ± 10.6, and 51.8 ± 1.6 nm, respectively. AgNPs obtained from different cellulose derivatives and various preparation parameters possess different inhibition potential against Escherichia coli and Staphylococcus aureus. The cellulose-AgNPs have higher effective against E. coli than S. aureus. HPMC-AgNPs showed significantly higher antibacterial activity than MC- AgNPs and HEC-AgNPs, respectively. These results suggest that the type of cellulose derivatives and the reaction parameters of the synthesis such as pH, temperature, and reaction period play an important role to the yield and physicochemical property of the obtained AgNPs.

Reduced ADAM8 levels upon non-surgical periodontal therapy in patients with chronic periodontitis.

OBJECTIVE: To determine effect of non-surgical periodontal treatment on a disintegrin and metalloproteinase 8 (ADAM8) levels in gingival crevicular fluid (GCF) of patients with chronic periodontitis (CP) in comparison with those of patients with gingivitis and to find correlations between ADAM8 levels and clinical parameters. DESIGN: Twenty-two and eleven patients with CP and gingivitis, respectively, were examined for four clinical parameters, probing depth, clinical attachment level, gingival and plaque indices. GCF from the selected gingivitis or periodontitis sites with distinct severities was sampled by Periopaper strips. The non-surgical treatments, including scaling and/or root planing and oral hygiene instruction, were provided for all patients. Clinical measurements and GCF sampling were repeated at three months after the treatments. ADAM8 concentrations were analyzed by ELISA and normalized by GCF volumes or total protein amounts. RESULTS: All patients exhibited significant improvement of almost every clinical parameter after treatment, whereas the median ADAM8 concentrations were significantly decreased at the moderate and severe periodontitis sites of patients with CP (p < 0.05). Moreover, the significantly positive correlations between ADAM8 concentrations and four clinical parameters were found in both moderate and severe groups (p < 0.05). CONCLUSION: ADAM8 concentrations were decreased by non-surgical periodontal therapy in patients with chronic periodontitis at the moderate and severe sites and were correlated with four clinical parameters, implying that GCF ADAM8 levels reflect inflammatory and bone-resorbing activities in the periodontal pocket.

Raised chondroitin sulphate WF6 epitope levels in gingival crevicular fluid in chronic periodontitis.

AIM: To determine the levels of chondroitin sulphate (CS) WF6 epitope, recognized by WF6 monoclonal antibody, in gingival crevicular fluid (GCF) from different stages of periodontal disease and healthy periodontium, and to correlate those levels with clinical parameters. MATERIAL AND METHODS: GCF samples, collected from 389 sites, were analysed for the WF6 epitope levels by the competitive enzyme-linked immunosorbent assay. RESULTS: The median WF6 epitope level was significantly higher in chronic periodontitis sites (n=185) than in healthy and gingivitis sites (n=204) (p<0.001), whereas the median levels did not significantly differ between healthy (n=65) and gingivitis sites (n=139). The median level in severe periodontitis sites (n=60) was significantly higher than that in moderate periodontitis sites (n=63) (p=0.019). Similarly, the median level in moderate periodontitis sites was significantly higher than that in slight periodontitis sites (n=62) (p=0.001). The WF6 epitope levels significantly correlated with probing depth (r=0.777, p=0.001) and loss of clinical attachment level (r=0.814, p=0.001). CONCLUSION: Elevated CS WF6 epitope levels in GCF are associated with severity of periodontitis. The WF6 antibody may therefore be clinically applied to monitor disease severity and progression.

Green synthesis and inhibitory effects against oral pathogens of silver nanoparticles mediated by rice extracts.

Rice is staple food for people in many countries for centuries. It is therefore considered as safe and environmental friendly material for pharmaceutical formulations. In the present study, aqueous extracts of three different parts of rice grain; rice bran (RB), rice husk (RH), and rice germ (RG) were compared for their use as reducing agents in synthesis of silver nanoparticles (AgNPs). AgNPs from those three different parts of rice, RB-AgNPs, RH-AgNPs, and RG-AgNPs, respectively showed different reducing activity, which the highest capacity was RB. RG-AgNPs and RB-AgNPs showed the maximum absorption of AgNPs at 440 nm whereas that of RH-AgNPs was at 480 nm. FTIR spectra of all AgNPs indicated the presence of different functional groups from rice attached to the nanoparticles and these groups prevented the particle agglomeration. Size analysis using dynamic light scattering revealed that RB-AgNPs was the smallest particles (346.4 ± 36.8 nm) and possessed the highest negative zeta potential. Antimicrobial test showed that the AgNPs obtained from green synthesis mediated by rice extracts have great antimicrobial activity against Streptococcus mutans, the severe oral pathogenic bacteria causing dental caries. These results suggest that aqueous extracts of RB, RH, and RG have potential to be used as reducing agents in synthesis of silver nanoparticles.

Efficacy of anesthetic rice nanogel on pain reduction in human oral cavity.

The aim of this study was to determine the efficacy of two local anesthetic rice nanogels (RNG) on pain reduction from needle insertion in oral cavity. Nanogel base was prepared using modified rice as gelling agent. The average particle size of RNG determined by photon correlation spectrophotometer was 485 ± 70 nm. Lidocaine hydrochloride (LH) and prilocaine hydrochloride (PH) were incorporated into RNG to obtain anesthetic RNG containing 5% and 20% LH or PH. Clinical efficacy test of each gel was performed in oral cavity of 100 healthy volunteers (25-60 years old). Evaluation was done by recording different pain measurements after inserting a needle into buccal mucosa after applying 5% and 20% anesthetic RNG. RNG base (placebo) and commercial anesthetic gels were used as negative and positive controls, respectively. It was found that the pain level in the negative control group was significantly higher than those of the anesthetic groups. Moreover, the pain level of the anesthetic RNG groups were lower than that of the commercial groups, especially in 20% anesthetic groups. For patient's satisfaction, most of the volunteers were appreciated with the anesthetic RNG as well as the commercial gels. They preferred to use high drug content RNG more than those with low drug content or placebo. It can be concluded that the anesthetic RNG has potential clinical efficacy in pain reduction during needle insertion in oral cavity.

Tooth whitening efficacy of pigmented rice gels containing carbamide peroxide.

Carbamide peroxide (CP) is commonly used as a tooth whitening agent. However, efficacy of CP can be enhanced if it is in the suitable delivery system. In the present study, CP loaded in pigmented rice gels were developed and investigated for their physicochemical properties and tooth whitening efficacy. The modified pigmented rice of two varieties, Homnil (HN) and Doisket (DS) were prepared and used as a gelling agent. The outer appearance of the obtained rice gels containing 10% CP (CP-HN and CP-DS, respectively) were transparent and homogeneous texture. The pH of both rice gel bases was neutral but became slightly acidic after incorporating with CP. The adhesive property of HN gel was significantly higher than DS gels. In vitro drug release profile exhibited that the release of CP from CP-DS was significantly higher than CP-HN and the commercial gel (CP-CG), respectively. In vitro tooth whitening efficacy in 45 normal teeth revealed that the tooth whitening efficacy of the gels was time dependent. At the end of the treatment, CP-HN showed significantly higher tooth whitening efficacy than CP-DS and CP-CG, respectively. It is concluded that the physicochemical properties, particularly the adhesive and dissolution properties, play an important role in the tooth whitening efficacy of the CP gels.

Effects of Piper betle fractionated extracts on inhibition of Streptococcus mutans and Streptococcus intermedius.

The overgrowth of certain strains of normal flora in oral cavity can cause many kinds of oral infections or diseases such as carries, periodontitis, and gingivitis. Prevention and treatment of these diseases are usually achieved by chemical antiseptics. However, these chemicals are found as negative impacts of human health hazards and accession of microbial resistance. The present study explores the potential of Piper betle extracts on inhibition of two oral pathogenic bacteria; Streptococcus mutans DMST 41283 and Streptococcus intermedius DMST 42700. P. betle demonstrated significantly higher inhibitory activity against both pathogenic strains than Acacia catechu, Camellia sinensis, Coccinia grandis, Solanum indicum, and Streblus asper. Among fractionated extracts of P. betle from several solvents, the extract from ethyl acetate (Pb-EtOAc) possessed the widest inhibition zone of 11.0 ± 0.1 and 11.3 ± 0.4 mm against both bacterial strains, respectively. Pb-EtOAc showed the same minimum inhibitory concentration of 0.5 mg/mL against both strains, whereas its minimum bactericidal concentrations were 2.0 and 0.5 mg/mL against S. mutans and S. intermedius, respectively. HPLC analysis demonstrated that the major active compound of Pb-EtOAc was 4-allylpyrocatechol. It was found that the killing kinetics of Pb-EtOAc against both test strains were time and dose dependent. Scanning electron microscopy micrographs showed the morphological changes and depletion of the tested pathogens indicating cell destruction after exposure to Pb-EtOAc. It is confirmed that Pb-EtOAc is potentially effective against both oral pathogens and might be used as natural alternative agents in prevention and treatment of oral infections caused by oral pathogenic bacteria.

Caesalpinia sappan: A promising natural source of antimicrobial agent for inhibition of cariogenic bacteria.

From the previous findings, the ethanolic fractionated extract of Caesalpinia sappan (F-EtOH) has high activity against Streptococcus mutans, the most severe cariogenic bacteria. The present study was aimed to isolate and identify the active compound of F-EtOH and compare its inhibitory activity against the biofilm of S. mutans as well as the cytotoxicity to oral fibroblast cells with F-EtOH. Compound isolation was done by column chromatography. The active compound was identified using liquid chromatography-mass spectrometry with electrospray ionization and nuclear magnetic resonance spectroscopy. It was found that the major compound of F-EtOH is brazilin. F-EtOH and brazilin were compared for inhibitory potential on the biofilms of three strains of S. mutans. The results exhibited that both F-EtOH and brazilin had potential on inhibiting biofilm formation and eradicating the preformed biofilms and their activity was dose dependent. F-EtOH showed significantly less toxic to normal periodontal ligament fibroblast than brazilin. At low concentration of 1- and 2-MBC, F-EtOH showed higher effective than brazilin. The results of our study suggest that the antibacterial activity of F-EtOH is according to the synergistic effects of the existing compounds including brazilin in F-EtOH.

Antioxidant activity and potential of Caesalpinia sappan aqueous extract on synthesis of silver nanoparticles.

The aim of this study was to investigate the antioxidant activity of Caesalpinia sappan aqueous extract (CE) and its potential on synthesis of silver nanoparticles (AgNPs). The antioxidant activity of CE was investigated using ferric reducing antioxidant power (FRAP) assay and two radical scavenging methods using 2,20-azinobis-(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS) and 1,1-diphenyl-2-picrylhydrazyl (DPPH) as free radicals. Silver nitrate (AgNO3) was used as precursor for the synthesis of AgNPs. Effects of AgNO3 concentration, reaction temperature, and duration of reaction were investigated. The obtained AgNPs was characterized using UV-Vis and photon correlation spectrophotometers. The antimicrobial activity of AgNPs was studied by means of diffusion method. The results from FRAP demonstrated that CE had high reducing property of 78.7 ± 2.4 mM Fe2+/mg. The trolox equivalent antioxidant capacity of CE determined by ABTS was 64.8 ± 4.2 µM/mg. The concentration of CE that can inhibit 50% of DPPH radicals (IC50) was 51.2 ± 3.2 µM. These results indicated that CE possesses strong antioxidant and reducing activities. The present study also showed that CE can act as reducing agent to produce AgNPs. The concentration of AgNO3, reaction temperature, and reaction time play an important role on the particles size and zeta potential of the obtained AgNPs. The antimicrobial activity of the AgNPs against Escherichia coli, Candida albicans, and Streptococcus mutants was stronger than against Staphylococcus aureus.