Evaluation of 1,9-Dimethyl-Methylene Blue nanoencapsulation using rhamnolipid nanoparticles to potentiate the Photodynamic Therapy technique in Candida albicans: In vitro study.

With the rapid development of nanotechnology, various functional nanomaterials have shown exciting potential in biomedical areas such as drug delivery, antitumor, and antibacterial therapy. These nanomaterials improve the stability and selectivity of loaded drugs, reduce drug-induced side effects, realize controlled and targeted drug release, and increase therapeutic efficacy. The increased resistance to antifungal microbicides in medical practice and their side effects stimulate interest in new therapies, such as Photodynamic Therapy (PDT), which do not generate resistance in microorganisms and effectively control the pathology. The present study aimed to evaluate, in vitro, the efficacy of photodynamic therapy on Candida albicans using 1,9-Dimethyl-Methylene Blue (DMMB) as photosensitizer, red LED (λ630), and nanoencapsulation of DMMB (RL-NPs/DMMB) using rhamnolipids produced by Pseudomonas aeruginosa to evaluate if there is better performance of DMMB + RL particles compared to DMMB alone via the characterization of DMMB + RL and colony forming count. The tests were carried out across six experimental groups (Control, DMMB, RL-NPs, RL-NPs/DMMB, PDT and PDT + RL-NPs/DMMB) using in the groups with nanoparticles, DMMB (750 ng/mL) encapsulated with rhamnolipids in a 1:1 ratio, the light source consisted of a prototype built with a set of red LEDs with an energy density of 20 J/cm2. The results showed that applying PDT combined with encapsulation (RL-NPs/DMMB) was a more practical approach to inhibit Candida albicans (2 log reduction) than conventional applications, with a possible clinical application protocol.

Photodynamic antimicrobial therapy (A-PDT) using 1,9-Dimethyl-Methylene Blue zinc chloride double salt - DMMB and λ640 ± 5ηm LED light in patients undertaking orthodontic treatment.

Orthodontic treatment involves the use of apparatuses that impairs oral hygiene making patients susceptible to periodontal diseases and caries. To prevent increased antimicrobial resistance A-PDT has shown itself a feasible option. The aim of this investigation was to assess the efficiency of A-PDT employing 1,9-Dimethyl-Methylene Blue zinc chloride double salt - DMMB as a photosensitizing agent combined with red LED irradiation (λ640 ± 5 ηm) against oral biofilm of patients undertaking orthodontic treatment. Twenty-one patients agreed to participate. Four biofilm collections were carried out on brackets and gingiva around inferior central incisors; first was carried out before any treatment (Control); second followed five minutes of pre-irradiation, the third was immediately after the first AmPDT, and the last after a second AmPDT. Then, a microbiological routine for microorganism growth was carried out and, after 24-h, CFU counting was performed. There was significant difference between all groups. No significant difference was seen between Control and Photosensitizer and AmpDT1 and AmPDT2 groups. Significant differences were observed between Control and AmPDT1 and AmPDT2 groups, Photosensitizer and AmPDT1 and AmPDT2 groups. It was concluded that double AmPDT using DMBB in nano concentration and red LED was capable to meaningfully decrease the number of CFUs in orthodontic patients.

Evaluation of dual application of photodynamic therapy-PDT in Candida albicans.

This study aimed to evaluate, in vitro, the efficacy of photodynamic therapy - PDT using dimethyl methylene blue zinc chloride double salt (DMMB) and red LED light on planktonic cultures of Candida albicans. The tests were performed using the ATCC 90,028 strain grown at 37 °C for 24 h, according to a growth curve of C. albicans. The colonies were resuspended in sterile saline adjusted to a concentration of 2 × 108 cells / mL, with three experimental protocols being tested (Protocol 1, 2 and 3) with a fixed concentration of 750 ɳg/mL obtained through the IC50, and energy density 20 J/cm2. Protocol 1 was carried out using conventional PDT, Protocol 2 was applied double PDT in a single session, and Protocol 3 was applied double PDT in two sessions with a 24 h interval. The results showed logarithmic reductions of 3 (4.252575 ± 0.068526) and 4 logs (2.669533 ± 0.058592) of total fungal load in protocols 3 and 2 respectively in comparison to the Control (6.633547 ± 0.065384). Our results indicated that double application in a single session of PDT was the most effective approach for inhibiting the proliferation of Candida albicans (99.991% inhibition).

Enhancement of photodynamic inactivation of planktonic cultures of Staphylococcus aureus by DMMB-AuNPs.

Photodynamic inactivation is a promising method for the treatment of infectious diseases. Nanotechnology through gold nanoparticles, as a tool to improve the delivery of photosensitizer is an attractive approach to enhance photodynamic inactivation of bacteria. Moreover, gold nanoparticles enchance the absorption of light due to their plasmon resonance. The aim of this study was to evaluate in vitro photodynamic inactivation effects of 1.9-Dimethyl-Methylene Blue (DMMB)-AuNPs associated with the red LED (λ630 ηm ± 20 ηm, 125 mW, 12 J / cm², 192 s) on S. aureus strain. Eight experimental groups were studied: Control, LED, AuNPs, AuNPs + LED, DMMB, DMMB + LED, DMMB + AuNPs, DMMB + AuNPs + LED. After incubation, the number of bacteria surviving each treatment was determined and then enumerated by viable counting (CFU / mL). The logarithm of CFU / mL (CFU/mL log10) was calculated. All experiments realized in triplicate. The statistical analyses included one-way ANOVA tests, Tukey's multiple comparisons and nonlinear regression, p values <0.05 were considered statistically significant. According to results, the photodynamic inactivation of S. aureus on groups DMMB + LED and DMMB-AuNPs + LED, showed a significant reduction of the microbial load (p < 0.0001) when compared to the Control group. The decimal reduction (RD) of these groups were 99.96 % (RD = 3) and 99.994 % (RD = 4) respectively. In conclusion, these findings demonstrated that photodynamic inactivation is enhanced by using DMMB-AuNPs on S. aureus.

A novel technique of antimicrobial photodynamic therapy - aPDT using 1,9-dimethyl-methylene blue zinc chloride double salt-DMMB and polarized light on Staphylococcus aureus.

Antimicrobial Photodynamic Therapy (aPDT) is an alternative to conventional treatments of local infections such as the use of antibiotics, which may lead to the development of resistance. aPDT besides requiring the use of a photosensitiser also needs a light source do be carried out. In the search for efficient and low-cost procedure the use of multispectral polarized light (λ400-2000 nm) emerges as a possibility for the execution of aPDT. The use of a highly effective photosensitizer is also of great importance. 1,9-Dimethyl-Methylene Blue Zinc Chloride Double Salt - DMMB is a potent phenothiazine derivative that presents high photodynamic action due to its high lipophilicity as well as a greater quantum yield of Singlet oxygen and phototoxicity when compared to other Photosensitizers. The aim of this study was to assess, In Vitro, the efficacy of aPDT on Staphylococcus aureus (ATCC 25923) using different concentrations of DMMB associated to a Polarized light source (Bioptron®, 40 mW, ᴓ = 15.8 cm2) using different energy densities. Based on the IC50, 150 and 300 ng/mL of DMMB concentrations were chosen for this study. Twelve experimental groups were used: (Control, PLs, PSs and aPDTs). Serial dilutions (up to 10-8) of the bacterial inoculum were used and the DMMB was added using the two previously determined concentrations. After 5 min of preincubation the dilutions of the inoculum were illuminated by the polarized light source. Subsequently, 100 µL of each dilution, in triplicate, were inoculated into Petri dishes containing TSA medium and incubated in a bacteriological oven at 37 °C for 24-h and quantification of UFCs was done. The results showed significant exponential reduction (p < .0001) of 99.93% (150 ng/mL + LP 10 J/cm2) and 99.97% (300 ng/mL + LP 5 J/cm2) the CFU counts in comparison to non-illuminated control. The results of this study allow to conclude that aPDT carried out with 1,9-Dimethyl-Methylene Blue Zinc Chloride Double Salt-DMMB and a PL souce was efficacious on the reduction (99.97%), in vitro, of the bacterial counts of S. aureus.

Oral microbiological control by photodynamic action in orthodontic patients.

BACKGROUND: Orthodontics involves diagnosis and treatment of dental and skeletal malocclusions. Orthodontic apparatus may repair these malocclusions but may also impair oral hygiene making patients prone to develop both periodontal diseases and caries. Antimicrobial agents may be used to prevent this.To avoid increased antimicrobial resistance to available drugs, A-PDT (Antimicrobial Photodynamic Therapy) appears as a viable alternative. OBJECTIVE: This work aimed to evaluate the efficacy of A-PDT on reducing the number of colony forming units (CFU) through the use of phenothiazine compound (methylene blue+ toluidine blue) as a photosensitizer, associated with red LED (λ640±5ηm) irradiation in orthodontic patients. METHODOLOGY: Twenty-one patients consented to participate in the study. Three biofilm collections were performed around the brackets and gums of the inferior central incisors; first before any intervention (Control); second after 5min of pre-irradiation and the last one immediately after AmPDT. Subsequently, a microbiological routine for microorganism growth period were performed and CFU counting after a 24h done. RESULTS: The data showed that the AmPDT was able to reduce CFU count around 90% when compared to Control group (p=0.007) and also between the A-PDT and Photosensitizer groups (p=0.010). However, there were no differences between the Control and Photosensitizer groups. CONCLUSION: A-PDT associated with the use of phenothiazine compounds and red LED was able to significantly reduce the number of CFUs in orthodontic patients.