Vaterite microparticle-loaded methylene blue for photodynamic activity in macrophages infected with Leishmania braziliensis.

Calcium carbonate (CaCO3) exhibits a variety of crystalline phases, including the anhydrous crystalline polymorphs calcite, aragonite, and vaterite. Developing porous calcium carbonate microparticles in the vaterite phase for the encapsulation of methylene blue (MB) as a photosensitizer (PS) for use in photodynamic therapy (PDT) was the goal of this investigation. Using an adsorption approach, the PS was integrated into the CaCO3 microparticles. The vaterite microparticles were characterized by scanning electron microscopy (SEM) and steady-state techniques. The trypan blue exclusion method was used to measure the biological activity of macrophages infected with Leishmania braziliensis in vitro. The vaterite microparticles produced are highly porous, non-aggregated, and uniform in size. After encapsulation, the MB-loaded microparticles kept their photophysical characteristics. The carriers that were captured allowed for dye localization inside the cells. The results obtained in this study indicated that the MB-loaded vaterite microparticles show promising photodynamic activity in macrophages infected with Leishmania braziliensis.

Gelatin nanoparticles via template polymerization for drug delivery system to photoprocess application in cells.

Photodynamic therapy (PDT) is a clinical treatment based on the activation of light-absorbing photosensitizers (PS) to generate reactive oxygen species, which are toxic to the targeted disease cells. Because most PS are hydrophobic with poor water solubility, it is necessary to encapsulate and solubilize PS in aqueous conditions to improve the photodynamic action for this compound. In this work, gelatin-poly(acrylic acid) nanoparticles (PAA/gelatin nanoparticles) via template polymerization for incorporation aluminum chloride phthalocyanine (ClAlPc) as a model drug for PDT application were developed. Biocompatible core-shell polymeric nanoparticles were fabricated via template polymerization using gelatin and acrylic acid as a reaction system. The nanoparticulate system was studied by scanning electron microscopy, steady-state, and their biological activity was evaluated using in vitro cancer cell lines by classical MTT assay. The obtained nanoparticles had a spherical shape and DLS particle size were determined further and was found to be around 170 nm. The phthalocyanine-loaded-nanoparticles maintained their photophysical behaviour after encapsulation. It is found that ClAlPc can be released from the nanoparticles in a sustained manner with a small initial burst release. In vitro cytotoxicity revealed that ClAlPc-loaded nanoparticles had similar cytotoxicity to free ClAlPc with mouse melanoma cancer cell line (B16-F10). In vitro photoeffects assay indicated that the nanoparticle formulation was superior in anticancer effect to free ClAlPc on mouse melanoma cancer cell line B16-F10. The results indicate that ClAlPc encapsulated in gelatin-poly(acrylic acid) nanoparticles are a successful delivery system for improving photodynamic activity in the target tissue.

Zinc phthalocyanine tetrasulfonate-loaded polyelectrolytic PLGA nanoparticles for photodynamic therapy applications.

BACKGROUND: Photodynamic Therapy (PDT) is a modality for the treatment of neoplastic tissues, which is based on the administration of a phototherapeutic agent and light irradiation at an appropriate wavelength, aiming to locate and destroy the target cell with the formation of reactive oxygen species. Nanoencapsulation technology presents itself as a tool for incorporation of bioactive substances aiming to improve their solubility in physiological environment, obtain a longer circulation time in the organism, administration of lower dosages and the minimization of side effects. The present work aimed at the development of poly (lactic acid-glycolic acid) (PLGA) nanoparticles coated with polyelectrolyte film layers for encapsulating zinc phthalocyanine tetrasulfonated (ZnPcSO4) as a bioactive substance model. METHODS: PLGA nanoparticles were produced by the double emulsion/solvent evaporation technique and polyelectrolytic coating was performed using polyalkylamine hydrochloride (PAH) as a weak polycation and poly (4-styrene sulfonate) (PSS) as a strong polyanion by layer-by-layer self-assembly technique (known as layer-by-layer-LbL). The nanoparticulate system was studied by scanning electron microscopy, steady-state, and their biological activity was evaluated using in vitro cancer cell lines by classical MTT assay. RESULTS: The polyelectrolytic PLGA nanoparticles had an average diameter of 384.7 ± 138.6 nm, restricted distribution size with a polydispersity index. The obvious change in zeta potential indicates successful alternation in polycation (PAH) and polyanion (PSS) deposition directly in PLGA nanoparticles. Scanning electron microscopy (SEM) analysis showed that the formed system had morphology spherical, typical of these release systems. The loading efficiency was 82.1 % ± 1.2 %. The polyelectrolytic nanoparticles loaded with phthalocyanine maintained their photophysical behavior after encapsulation. Cell viability was determined, obtaining 90 % cell death. CONCLUSIONS: Therefore, the presented work depicts ZnPcSO4-loaded polyelectrolytic PLGA nanoparticles as a promise drug delivery system for phototherapeutic agent, which are thus expected to have superior therapeutic efficiency than free drug.

BSA nanoparticles loaded-methylene blue for photodynamic antimicrobial chemotherapy (PACT): effect on both growth and biofilm formation by Candida albicans.

It has been demonstrated an increase in resistance of Candida albicans to conventional therapies, probably, due the indiscriminate use of the conventional antifungal drugs. In this aspect, the nanotechnology generates the possibility of creating new therapeutic agents. Thus, the objective of this paper was to produce and characterize a bovine serum albumin (BSA) nanoparticle encapsulated with Methylene Blue (MB). In addition, the effect of BSA nanoparticles encapsulated with MB (BSA-MB) was evaluated on both growth and biofilm formation by C. albicans by Photodynamic Antimicrobial Chemotherapy (PACT) protocols. The BSA-MB nanoparticles were prepared by the desolvation process. The nanoparticulate system was studied by steady-state techniques, scanning electron microscopy and their biological activity was evaluated in vitro both growth and biofilm formation by C. albicans. The synthetized BSA-MB nanoparticles were spherical in shape exhibiting a 100-200 nm diameter with a low tendency to aggregate (PDI values < 0.2). MB photophysical properties were shown to be preserved after BSA encapsulation. A significant reduction in C. albicans growth, after PACT was observed, in a dependent manner on MB-loaded in BSA nanoparticles concentration used. It was observed an inhibition of 23, 65 and 83% in the presence of MB-loaded in BSA nanoparticles 0.1, 0.5 and 1.0 µg.mL-1, respectively. In addition, MB-loaded BSA nanoparticles 0.5 µg.mL-1 were able to reduce both biofilm formation (80%) and the transition from yeast to filamentous form by C. albicans. The results presented here demonstrated a potentiation of the phototoxic effect of MB after BSA encapsulation, since the concentrations of MB-loaded BSA nanoparticles necessary to inhibits ∼50% of C. albicans development was 10 times minor than that observed for free MB. Taken together, these results suggest the potential of PACT, using MB-loaded BSA nanoparticles in inhibiting C. albicans development. The synthesis and design of BSA nanoparticles can be successfully applied for MB encapsulation and offer the possibility to drive the toxicity effect to a specific target, as an evaluation on both growth and biofilm formation by Candida albicans.

Synthesis, photophysical and photobiological characterization of BSA nanoparticles loaded with chloroaluminium phthalocyanine by one-step desolvation technique for photodynamic therapy action.

This work describes the preparation of bovine serum albumin (BSA) nanoparticles by one-step desolvation method using acetone/ethanol as precipitating agent, glutaraldehyde as crosslinking agent, sodium azide as a preservative and water as reaction media for chloroaluminium phthalocyanine (ClAlPc) incorporation for photodynamic therapy action. The characterization of the nanoparticulate system was carried out using steady-state technique, scanning electron microscopy study and their biological activity was evaluated using in vitro macrophages cell lines by classical MTT assay. All the spectroscopy measurements demonstrated good photophysical properties and the in vitro cytotoxicity study showed that the system is not cytotoxic in the darkness, but it exhibits a substantial phototoxicity at 0.90 mol.L-1 of photosensitizer concentration and 10.0 J cm-2 of light. These conditions are sufficient to kill about 90% of the cells. The results allowed us to conclude that ClAlPc-loaded in BSA nanoparticles might have potential application on drug delivery system for PDT protocols.

Gelatin nanoparticles loaded methylene blue as a candidate for photodynamic antimicrobial chemotherapy applications in Candida albicans growth.

Gelatin nanoparticles (GN) with an intrinsic antimicrobial activity maybe a good choice to improve the effectiveness of photodynamic antimicrobial chemotherapy (PACT). The aim of this study was to development gelatin nanoparticles loaded methylene blue (GN-MB) and investigate the effect of GN-MB in the Candida albicans growth by PACT protocols. The GN and GN-MB were prepared by two-step desolvation. The nanoparticulate systems were studied by scanning electron microscopy and steady-state techniques, the in vitro drug release was investigated, and we studied the effect of PACT on C. albicans growth. Satisfactory yields and encapsulation efficiency of GN-MB were obtained (yield = 76.0% ± 2.1 and EE = 84.0% ± 1.3). All the spectroscopic results presented here showed excellent photophysical parameters of the studied drug. Entrapment of MB in GN significantly prolongs it's in vitro release. The results of PACT experiments clearly demonstrated that the photosensitivity of C. albicans was higher when GN-MB was used. Gelatin nanoparticles loaded methylene blue-mediated photodynamic antimicrobial chemotherapy may be used against Candida albicans growth.

Preparation of gelatin nanoparticles by two step desolvation method for application in photodynamic therapy.

Gelatin nanoparticles have recently been receiving considerable attention because they offer a good option as release systems due to their low cost, biocompatibility, biodegradability and its application in several types of formulations. This study aim was to evaluate the potential application of gelatin nanoparticles entrapping a photosensitizer in Photodynamic Therapy. Gelatin nanoparticles were studied by steady-state techniques and the biological activity evaluated by in vitro MTT assay. The particles were spherical in shape exhibiting a 273 nm diameter with a low tendency to aggregate. The loading efficiency was 76%. Photosensitizer photophysical properties were shown to be preserved after GN encapsulation. The cells viability obtaining 85% cells death compared with control. The results demonstrate that gelatin nanoparticles can be successfully applied for photosensitizers encapsulation or other active drugs and be used as an optimal medium for a variety of bioactive materials, which can also be encapsulated by the proposed method.

EFICÁCIA DA FONOFORESE COM XIMENIA AMERICANA L. NA INFLAMAÇÃO DE TENDÃO DE RATOS / PHONOPHORESIS EFFECTIVENESS WITH XIMENIA AMERICANA L. IN RATS TENDON INFLAMMATION / EFICACIA DE LA FONOFORESIS CON XIMENIA AMERICANA L. EN LA INFLAMACIÓN DE TENDÓN DE RATAS

RESUMO Introdução: Diversos recursos terapêuticos, como laser e ultrassom isolado, combinados com fármacos e fonoforese têm sido utilizados em casos de inflamação e reparo de tendão, sendo o ultrassom pulsado bastante eficaz em tendinites crônicas. Objetivo: Analisar a eficácia do uso da fonoforese com o extrato etanólico das cascas do caule da Ximenia americana L. na resolução do processo inflamatório crônico em Rattus norvegicus. Métodos: A amostra consistiu em 120 animais para análise da resposta inflamatória, utilizando-se como variáveis edema, força biomecânica, número de fibroblastos e análise histológica. Resultados: Nos 7° e no 14° dia, verificou-se diferença significativa entre o grupo ultrassom com gel do caule de Ximenia americana L. e o grupo ultrassom com placebo (p < 0,05) quanto à redução de edema, aumento da força máxima de ruptura, redução extremamente significativa da deformação máxima (p < 0,001), além de aumento dos fibroblastos. Na análise histológica, houve melhora do processo inflamatório inicial e aceleração do reparo tendíneo, com redução de células inflamatórias e com deposição de colágeno organizado com matriz extracelular densa. Conclusão: O ultrassom pulsado combinado com o gel do caule de Ximenia americana L. é uma forma terapêutica eficaz para a resolução do processo inflamatório crônico.

ABSTRACT Introduction: Several therapeutic resources such as laser and isolated ultrasound combined with drugs and phonophoresis have been used in cases of inflammation and tendon repair, and the pulsed ultrasound is quite effective in chronic tendinitis. Objective: To analyze the efficacy of phonophoresis with the ethanol extract of the stem bark of Ximenia americana L. to manage the chronic inflammatory process in Rattus norvegicus. Methods: The sample consisted of 120 animals for analysis of the inflammatory response using edema, biomechanical strength, number of fibroblasts and histological analysis as variables. Results: On the 7th and 14th day, there was a significant difference between the group ultrasound with Ximenia americana L. gel and the ultrasound with placebo group (p<0.05) for edema reduction, increased maximum rupture strength, highly significant reduction of the maximum deformation (p<0.001), in addition to an increase in fibroblasts. In the histological analysis, there was improvement in the inflammatory process and acceleration of tendon repair with reduction of inflammatory cells and deposition of organized collagen with dense extracellular matrix. Conclusion: The pulsed ultrasound combined with gel of Ximenia americana L. is an effective therapy to manage the chronic inflammatory process.

RESUMEN Introducción: Varios recursos terapéuticos, tales como láser y ultrasonido aislado, en combinación con fármacos y la fonoforesis se han utilizado en casos de inflamación y reparación del tendón, y el ultrasonido pulsado ha sido muy eficaz en la tendinitis crónica. Objetivo: Analizar la eficacia de la fonoforesis con el extracto de etanol de la corteza del tallo de Ximenia americana L. en la resolución del proceso inflamatorio crónico en Rattus norvegicus. Métodos: La muestra fue de 120 animales para analizar la respuesta inflamatoria, utilizando como variables el edema, la resistencia biomecánica, el número de fibroblastos y el análisis histológico. Resultados: En el 7° y en el 14° día, se observó una diferencia significativa entre el grupo de ultrasonido con gel de Ximenia americana L. y el grupo ultrasonido con placebo (p < 0,05) en la reducción del edema, el aumento de la resistencia máxima a la rotura, la reducción altamente significativa de la deformación máxima (p < 0,001), además del aumento de los fibroblastos. En el análisis histológico hubo una mejoría en el proceso inflamatorio y la aceleración de la reparación del tendón, con reducción de células inflamatorias y deposición de colágeno organizado con matriz extracelular densa. Conclusión: La eficacia del ultrasonido pulsado combinado con gel de Ximenia americana L. es una forma eficaz para la resolución del proceso inflamatorio crónico.

Comparison of the effect of rose bengal- and eosin Y-mediated photodynamic inactivation on planktonic cells and biofilms of Candida albicans.

Candida albicans is an opportunistic yeast that can cause oral candidosis through the formation of a biofilm, an important virulence factor that compromises the action of antifungal agents. The objective of this study was to compare the effect of rose bengal (RB)- and eosin Y (EY)-mediated photodynamic inactivation (PDI) using a green light-emitting diode (LED; 532 ± 10 nm) on planktonic cells and biofilms of C. albicans (ATCC 18804). Planktonic cultures were treated with photosensitizers at concentrations ranging from 0.78 to 400 µM, and biofilms were treated with 200 µM of photosensitizers. The number of colony-forming unit per milliliter (CFU/mL) was compared by analysis of variance and Tukey's test (P ≤ 0.05). After treatment, one biofilm specimen of the control and PDI groups were examined by scanning electron microscopy. The photosensitizers (6.25, 25, 50, 200, and 400 µM of EY, and 6.25 µM of RB or higher) significantly reduced the number of CFU/mL in the PDI groups when compared to the control group. With respect to biofilm formation, RB- and EY-mediated PDI promoted reductions of 0.22 log10 and 0.45 log10, respectively. Scanning electron microscopy showed that the two photosensitizers reduced fungal structures. In conclusion, EY- and RB-mediated PDI using LED irradiation significantly reduced C. albicans planktonic cells and biofilms.

Susceptibility of planktonic cultures of Streptococcus mutans to photodynamic therapy with a light-emitting diode.

The objective of this study was to evaluate the effect of photodynamic therapy with erythrosine and rose bengal using a light-emitting diode (LED) on planktonic cultures of S. mutans. Ten S. mutans strains, including nine clinical strains and one reference strain (ATCC 35688), were used. Suspensions containing 106 cells/mL were prepared for each strain and were tested under different experimental conditions: a) LED irradiation in the presence of rose bengal as a photosensitizer (RB+L+); b) LED irradiation in the presence of erythrosine as a photosensitizer (E+L+); c) LED irradiation only (P-L+); d) treatment with rose bengal only (RB+L-); e) treatment with erythrosine only (E+L-); and f) no LED irradiation or photosensitizer treatment, which served as a control group (P-L-). After treatment, the strains were seeded onto BHI agar for determination of the number of colony-forming units (CFU/mL). The results were submitted to analysis of variance and the Tukey test (p ≤ 0.05). The number of CFU/mL was significantly lower in the groups submitted to photodynamic therapy (RB+L+ and E+L+) compared to control (P-L-), with a reduction of 6.86 log10 in the RB+L+ group and of 5.16 log10 in the E+L+ group. Photodynamic therapy with rose bengal and erythrosine exerted an antimicrobial effect on all S. mutans strains studied.

Susceptibility of planktonic cultures of Streptococcus mutans to photodynamic therapy with a light-emitting diode

The objective of this study was to evaluate the effect of photodynamic therapy with erythrosine and rose bengal using a light-emitting diode (LED) on planktonic cultures of S. mutans. Ten S. mutans strains, including nine clinical strains and one reference strain (ATCC 35688), were used. Suspensions containing 10(6) cells/mL were prepared for each strain and were tested under different experimental conditions: a) LED irradiation in the presence of rose bengal as a photosensitizer (RB+L+); b) LED irradiation in the presence of erythrosine as a photosensitizer (E+L+); c) LED irradiation only (P-L+); d) treatment with rose bengal only (RB+L-); e) treatment with erythrosine only (E+L-); and f) no LED irradiation or photosensitizer treatment, which served as a control group (P-L-). After treatment, the strains were seeded onto BHI agar for determination of the number of colony-forming units (CFU/mL). The results were submitted to analysis of variance and the Tukey test (p < 0.05). The number of CFU/mL was significantly lower in the groups submitted to photodynamic therapy (RB+L+ and E+L+) compared to control (P-L-), with a reduction of 6.86 log10 in the RB+L+ group and of 5.16 log10 in the E+L+ group. Photodynamic therapy with rose bengal and erythrosine exerted an antimicrobial effect on all S. mutans strains studied.

Atividade antifúngica do óleo essencial de Cymbopogon citratus frente a cepas de Candida albicans e Candida tropicalis isoladas de infecções nosocomiais / Antifungal activity of Cymbopogon citratus essential oil on Candida albicans and Candida tropicalis strains isolated from nosocomial infections

O óleo essencial de Cymbopogon citratus tem sido alvo de vários estudos em função do potencial antimicrobiano. Neste estudo, a atividade desse componente foi investigada em cepas do gênero Candida isoladas de infecções hospitalares. Para a condução do estudo, foram analisadas 24 isolados de Candida albicans e 15 isolados de Candida tropicalis, originados de pacientes com suspeitas de infecção hospitalar e uma cepa padrão de C. albicans ATCC 10231, por meio da técnica de difusão em ágar. O óleo essencial de C. citratus apresentou atividade antifúngica em 100% dos isolados a partir da concentração de 25% (v/v), o que indica sua ação positiva sobre as cepas hospitalares. Sugere-se a realização de estudos farmacológicos e toxicológicos desse componente para avaliar a possível aplicação clínica.

Atividade antifúngica do citral em leveduras do gênero Candida isoladas de pacientes hospitalizados / Citral antifungal activity against Candida genus yeasts isolated from hospitalized patients

O citral, componente do óleo essencial de Cymbopogon citratus, tem sido pesquisado para verificar sua ação antimicrobiana em bactérias e fungos. Com o objetivo de avaliar a atividade antifúngica do citral contra leveduras do gênero Candida, no presente trabalho foram avaliadas 32 amostras de Candida albicans, 25 de C. tropicalis, 20 de C. parapsilosis e 5 de C. glabrata, coletadas de pacientes hospitalizados. O citral foi testado nas concentrações de 10%, 15%, 25%, 35%, 50% e 60% (v/v), utilizando a técnica de difusão em ágar Sabouraud. A atividade antifúngica do citral foi constatada em todas as leveduras selecionadas nas concentrações ≥25%. Mediante os resultados obtidos, sugere-se a realização de novas pesquisas sobre citral frente às demais espécies de fungos patogênicos para conhecer as características toxicológicas e farmacológicas para que esse componente possa futuramente ser utilizado como um importante princípio ativo na produção de novos agentes antifúngicos.

Cytotoxicity of octal-bromide zinc phthalocyanine after photodynamic therapy with different light sources.

OBJECTIVE: The objective of this study was to investigate the cytotoxicity of octal-bromide zinc phthalocyanine (ZnPcBr8) at different concentrations (0.25, 0.5, and 1 microM) after irradiating HEp-2 cell cultures with two different light sources: a diode semiconductor laser (660 nm, 30 mW) or an LED (640 nm, 70 mW). In order to obtain comparative results, the irradiation parameters of both light sources were adjusted so that the amount of energy density delivered would be the same (4.5 J/cm2). BACKGROUND DATA: Numerous photosensitizers and light sources used in the treatment of human disease have been studied. Based on these studies, a comparative evaluation of two light sources used in photodynamic therapy (PDT) with ZnPcBr8 was proposed. MATERIALS AND METHODS: HEp-2 cells were incubated with ZnPcBr8 at different concentrations (0.25, 0.5, or 1 microM) for 1 h, irradiated with the diode semiconductor laser (660 nm at 30 mW for 300 sec; 4.5 J/cm2) or the LED laser (640 nm at 70 mW for 128 sec; 4.5 J/cm2), and then incubated in MEM medium for 1 or 24 h. The cells were analyzed using the MTT and trypan blue dye exclusion tests. RESULTS: The results demonstrated that the concentration of 1 microM of ZnPcBr8 was the most effective after PDT administered by both light sources. According to the MTT results, HEp-2-cell viability decreased by 97.96% 1 h after, and by 99.87% 24 h after irradiation with the diode semiconductor laser, and decreased by 94.03% 1 h after, and by 99.21% 24 h after irradiation with the LED. The results obtained using the trypan blue dye exclusion test confirmed the photodynamic efficacy of ZnPcBr8 employed with both light sources. With regard to HEp-2-cell viability, the following results were observed: a decrease of 98.73% 1 h after, and of 99.49% 24 h after irradiation with the diode semiconductor laser; and a decrease of 98.76% 1 h after, and of 99.23% 24 h after irradiation with the LED. CONCLUSIONS: According to our results with the irradiation parameters studied here, both the LED and diode semiconductor laser can be used for PDT in vitro, since both light sources had excellent photodynamic efficacy.