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OBJECTIVE: To study the tissue distribution differences of chrysophanol loaded polybutylcyanoacrylate nanocapsules, ehrysophanol-hydroxypropyl-β-cyclodextrin inclusion complex, chrysophanol liposomes in mice, through the tissue distribution of chrysophanol formulations to understand the pharmacodynamic properties of the formulations. With a view to electing for brain targeting the most strong, effect of chrysophanol best formulation. METHODS: Mice tail intravenous injection of the equivalent dose of 10 mg · kg-1 chrysophanol formulations, by 0.25, 1, 2, 4, 8, 12 h taking different points of mice heart, liver, spleen, lungs, kidneys, brain and blood plasma, after playing homogenized the protein processing using HPLC for the determination of chrysophanol in organizations of distribution. Chrysophanol was analyzed on a Thermo Hypersil ODS2 chromatographic column with mobile phase methanol-water(90:10) at 1.0 mL · min-1 flow rate and with column temperature 30°C. The wavelength of UV detector was set at 254 nm and injection volume was 20 μL. RESULTS: Three kinds of chrysophanol formulations could prolong the elimination of time in vivo, and better targeting of tissues and organs than chrysophanol N, N-DMF solution. CONCLUSION: Three kinds of chrysophanol formulations compared to tissue distribution, chrysophanol liposomes in various tissue distribution better, brain tissue targeting the most significant. Chrysophanol liposomes is expected to become the treatment of cerebrovascular disease clinical application of new dosage forms.
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Materiais poliméricos têm sido usados em um vasto número de aplicações médicas e farmacêuticas, caracterizando-se como biomateriais. Antes do uso humano, se faz necessário uma criteriosa programação de testes que permitam investigação da segurança do material e avaliação da eficácia e assim, avaliar sua biocompatibilidade. Essa atenção também tem sido aplicada a materiais nanoparticulados, inclusive de constituição polimérica. Sendo assim, antes da liberação destes materiais para uso humano, testes preliminares são feitos in vitro e in vivo, neste caso com experimentação animal. Paralelamente à crescente busca em se manter a segurança biológica, a ciência tem procurado metodologias alternativas para se obter resultados seguros e que possam substituir os obtidos in vivo. Neste contexto, surge grande interesse no uso de culturas celulares, primeiramente em monocamadas e posteriormente, o desenvolvimento de culturas celulares em estrutura tridimensional, mimetizando de maneira mais próxima o que ocorre in vivo. Considerando as diversas vantagens obtidas no estudo de culturas tridimensionais, este estudo teve como objetivo desenvolver tal modelo, visando simular sistemas de avaliação para biocompatibilidade de materiais poliméricos, testando nanopartículas de polibutilcianoacrilato. Foram cultivadas células de fibroblasto e melanócito humanos e células de melanoma (SK-Mel-103) em monocamada e em matriz tridimensional de colágeno tipo I e avaliada a citotoxicidade em 24, 48 e 72 horas das nanopartículas utilizando os testes de Exclusão Azul de Tripan e MTT. Os resultados indicam que as nanopartículas de polibutilcianoacrilato são mais citotóxicas com o aumento da concentração da amostra, assim como o IC50 foi decrescente ao longo dos dias, demonstrando a intensificação da resposta biológica no decorrer do tempo e foram mais citotóxicas para as células de melanoma. Este perfil de aumento de citotoxicidade com o passar do tempo também foi observado no ensaio clonogênico, onde as células foram cultivadas em monocamada e incubadas com a amostra até 14 dias. Os testes de viabilidade celular em cultura tridimensional evidenciaram faixa de IC50 um pouco maior do que em monocamada, demonstrando uma pequena redução na citotoxicidade quando avaliada no ambiente 3D. Foram feitos testes de caracterização do tipo de morte utilizando citometria de fluxo, confirmando os dados dos testes de viabilidade que indicam aumento de morte com o aumento da concentração de teste. A morte acontece por apoptose tardia. Nas concentrações próximas ao IC50, há indicação que as nanopartículas de polibutilcianoacrilato inibem a autofagia em fibroblastos e células de melanomas cultivados em monocamada, característica que é anulada quando o comportamento é avaliado em ambiente tridimensional. Os resultados indicam que nanopartículas de polibutilcianoacrilato podem levar a modificação da resposta celular dependendo da concentração empregada e do ambiente de cultivo das células, mostrando serem promissores o estudo e o emprego de ambiente tridimensional na avaliação de citotoxicidade de nanopartículas poliméricas
Polymeric materials have been widely used as biomaterials in medical and pharmaceutical applications. Before their use in humans, a number of tests is necessary to investigate the safety and effectiveness of the biomaterial in order to determine its biocompatibility. This attention has been given to nanoparticulate materials, including those of polymeric composition. Thus, in vitro and in vivo preliminary tests are done before these materials are released for human use. In parallel with the concern with biological safety studies, there has been a continuous effort in finding alternative methods to replace the current in vivo animal tests and provide reliable results regarding safety. In this context, great interest arises in the use of cell culture, firstly as monolayer and secondly as three-dimensional cell culture systems that mimic more closely the in vivo morphology. Taking into account all the advantages of three-dimensional cell culture studies, the aim of this work is to develop a three-dimensional model that can effectively evaluate polymeric biomaterials biocompatibility using polybutylcyanoacrylate nanoparticles. Cell culture of human fibroblasts, melanocytes and melanoma cells (SK-Mel-103) were cultured in monolayer and on type I collagen matrix. Polybutylcyanoacrylate nanoparticles cytotoxicity was evaluated in 24, 48 and 72 hours using Tripan Blue and MTT assays and the results found were dose and time-dependent to all cell types. Moreover, melanoma cells were significantly more sensitive to the nanoparticles toxicity. The same profile of cytotoxic response was observed in the clonogenic assay, where cells were cultured in low-density monolayer and incubated with the nanoparticles for 14 days. Cell viability assays in 3D culture had a slightly higher IC50 range when compared to the monolayer results, which suggests a reduction in the nanoparticle cytotoxicity in the 3D environment. Cell death analysis using flow cytometry confirmed the dose-dependent response obtained by cell viability assays. Cell death occurred mostly via late apoptosis. There is suggestive evidence that treatment with polybutylcyanoacrylate nanoparticles in concentrations close to the IC50 inhibits autophagy in fibroblasts and melanoma cells when cultured in monolayers, but this response could not be observed in the 3D environment. Our results showed that polybutylcyanoacrylate nanoparticles can modify cell response depending on the concentration used and the conditions of cell culture. We conclude that the 3D cell culture is a promising method for the evaluation of polymeric nanoparticles cytotoxicity
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Animals , Cells , Biocompatible Materials , Materials Testing , Cell Culture Techniques , Imaging, Three-Dimensional , NanoparticlesABSTRACT
Objective: Optimizing the formula and preparation of β-elemene- polybutylcyanoacrylate nanoparticles (β-ELE-PBCA-NP). Methods: With polybutylcyanoacrylate (PBCA) as the drug-loaded material, β-ELE-PBCA-NP was prepared by interfacial polycondensation. The preparation conditions were optimized by single factor design and the formula was optimized by orthogonal design with entrapment efficiency (EE) as index. EE of β-ELE-PBCA-NP was determinated by RP-HPLC. Results: Under the optimal conditions, the prepared NPs were round and regular in shape without adhesions with average particle size of 254 nm and EE of 90.17%. Conclusion: The optimized technology in this experiment is stable and feasible.
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Objective: To prepare amphotericin B-polybutylcyanoacrylate-loaded nanoparticle (AmB-PBCA-NP) and to assess their ability to pass the blood-brain barrier. Methods: AmB-PBCA-NP was prepared by incubation, with the surface modified by polysorbate 80. High-performance liquid chromatography (HPLC) method was established using acetonitrile and water (40:60, 4% of acetic acid) as mobile phase; detection was done at 405 nm. Mice were divided into 3 groups: AmB-injected, AmB-lipid-injected, and AmB-PBCA-NP-injected group. Brain-targeting ability of AmB-PBCA-NP was then evaluated by determination of AmB concentrations in mice brain and other specimens. Results: The mean diameter of the prepared AmB-PBCA-NP was 94.38 nm; the mean entrapment efficiency of the particles was 82%; and the drug loading rate was 56.10%. It was found that AmB alone failed to pass the blood-brain barrier; only very low level of AmB was detected in AmB-lipid group 3 h after injection; moderate concentration of Amb was detected 30 min after injection of AmB-PBCA-NP and the concentration peaked at 133 ng/g 3 h after injection. Conclusion: AmB-PBCA-NP modified by polysorbate 80 can pass the blood-brain barrier.
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OBJECTIVE: To prepare Mitomycin C for injection-Polybutylcyanoacrylate Nanoparticles (MMC-PBCA-NPs) freeze drying preparation by microfluidization and to investigate its hemolytic feature. METHODS: The crude emulsion of MMC-PBCA-NPs was prepared by emulsion polymerization method, from which MMC-PBCA-NPs solution was prepared by microfluidization method; the encapsulation efficiency (EE) and the drug loading (DD) of the preparation were detected by ultraviolet spectrophotometry. The particle size and appearances of which were observed. A hemolytic test was performed on rabbits to observe the blood reaction of the preparation. RESULTS: The prepared MMC-PBCA-NPs freeze drying preparation was uniformly distributed and in round shape, with EE, DD and mean diameter at (85.1?3.8)%, (7.0?0.2)% and (113.5?3.86)nm, respectively. The hemolytic test was negative. CONCLUSION: It is feasible to prepare MMC-PBCA-NPs by microfluidization method.
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OBJECTIVE:To optimize the formula and preparation technique of oleanolic acid loaded polybutylcyanoacrylate nanocapsules(OA-PBCA-NC)by interfacial polymerization and to study its quality.METHODS:The preparation technique of OA-PBCA-NC prepared by interfacial polymerization were optimized by single factor design,and the formula was optimized by orthogonal design with entrapment ratio as index.Its shape,particle size and the entrapment ratio were investigated as well.RESULTS:The optimized dosage ratios were as follows∶ OA vs.BCA∶60mg∶0.1ml;acetic ether vs.BCA∶2.0ml∶0.1ml;and oil vs.water:1∶1.The prepared nanocapsules were round and regular in shape without adherence.The particle size was well-distributed with mean diameter at(214?8)nm and mean entrapment ratio at(76.8?0.4)%.CONCLUSION:The preparation technique of OA-PBCA-NC established in this study is stable and feasible.
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Objective:To prepare amphotericin B(AmB)-polybutylcyanoacrylate(PBCA) nanoparticle and evaluate its characteristics,so as to screen for the optimal preparation technique.Methods: AmB was loaded to blank PBCA nanoparticles by incubation to prepare AmB-PBCA nanoparticles.The surface of the nanoparticles was modified with polysorbate 80.The AmB concentration of AmB-PBCA-NP solution was calculated with the standard curve of AmB at D_(405),and the stability of solution was assessed.The diameter distribution of the particles was determined with Coulter Laser granulometer.The colloid solution of AmB-PBCA-NP was centrifuged and the supernatant was harvested to determine the concentration of AmB.The encapsulation rate,drug loading rate,and in vitro drug release were measured.The optimized preparation recipe was screened using diameter,drug loading rate,and encapsulation rate of the particles as indices.Results: The prepared AmB-PBCA nanoparticle had a regular spherical or elliptic surface,with a mean diameter of(69.01?28.56) nm.The curve of standard AmB was linear within the range of 1.12-5.60 ?g/ml: D_(405)=0.163 4c+0.006 6(r=0.999 3).The average recovery of AmB-PBCA-NP was(99.93%,) showing the solution of AmB-PBCA-NP was stable within 12 h.The nanoparticles showed a sustained drug release in vitro within 24 h.The optimized recipe was: DextranT-70 stabilizer without sodium deoxycholate,with a mean encapsulation rate of 56.10% and a drug loading rate of 82%.Conclusion: The current method for preparing AmB-PBCA nanoparticles is simple and can meet the requirements of pharmaceutics.
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Objective To prepare magnetic polybutylcyanoacrylate (PBCA) nanosphere loaded with aclacinomycin A (ACM) and investigate its anti carcinoma effect and toxicity. Methods Magnetic PBCA nanospheres loaded with ACM were prepared by interfacial polymerization method. Female BABL/c nude mice were injected with MKN 45 gastric carcinoma cell line mass subcutaneously near right forefoot to establish human gastric carcinoma model. The mice were divided into 5 groups with 6 mice in each group: ACM (8 mg/kg), nanosphere with low dosage of ACM(1.6 mg/kg), nanosphere with high dosage of ACM (8 mg/kg ), magnetic nanosphere without ACM and normal saline (NS). Magnets (2500 Gs) were embedded in the tumor mass of all the mice one day before the date of therapy. The agents were administered again the same as the first time after 5 days. Tumor weight was measured and assay of colony forming unit granulocyte and macrophage (CFU GM) was performed on semi solid culture. Results The drug content of the agent was 12.0% and the mean particle size was 210 nm. The tumor inhibition rates of ACM, nanosphere with low dosage of ACM, nanosphere with high dosage of ACM and carrier without ACM on human gastric carcinoma in nude mice were 22.63%, 30.66%, 52.55% and 10.22% respectively. The average tumor weight of targeted nanospheres group was much lower than that of the same dosage of non targeted ACM group( P
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Objective To optimize the preparation technology of chrysophanol loaded polybutylcyanoacrylate nanocapsules by interfacial polymerization,sieve the optimal technology conditions of preparation,and study its quality. Methods Based on the single factor experiment,the formulation was optimized by L9(34) orthogonal design with entrapment efficiency as reference index. The factors including agitating speed,pH value of aqueous phase,amount of ?-butylcyanoacrylate (BCA) and of ethyl acetate were screened. At last,its shape,particle size,the encapsulation efficiency,and the drug loading were investigated as its quality inspection. Results When the ammunt of chrysophanol was 5 mg,the optimum preparation conditions were established as follows:the agitating speed was 800 r/min,the pH value of aqueous phase was 2,the amount of ?-BCA was 13 ?L and ethyl acetate was 0.6 mL. Under this optimum condition,the mean entrapment ratio was 82.19%,and the mean drug loading was 21.48%,the mean diameter of the nanocapsule was 246 nm. The particle size was well-distributed showed by electron microscope pictures. Conclusion The prepared chrysophanol loaded polybutylcy-anoacrylate nanocapsules has small size,high encapsulation efficiency,and drug loading. The preparation technique established in this study is stable and feasible. It also can be used for iv injection.