Optimized Chitosan-Coated Gliadin Nanoparticles Improved the Hesperidin Cytotoxicity over Tumor Cells

Abstract Hesperidin is a natural compound which is found in citric fruits and presents antitumor and antimicrobial activities. However, the in vivo efficacy of Hesperidin is reduced due to its low oral bioavailability. Protein-based nanoparticles have been applied to improve biological parameters of drugs and natural compounds. Gliadin is a monomeric protein present in wheat. In this study, gliadin-based nanoparticles containing hesperidin were obtained by desolvation technique and a Taguchi orthogonal array design was employed to optimize the formulation. The independent variables were set as concentration of CaCl2 (0.5; 1 or 2%) and stabilizing agent (Pluronic F68, Tween 80 or sodium caseinate). The dependent variables consisted of mean diameter, polydispersity index, zeta potential, and encapsulation efficiency. The results showed significant effects on the dependent variables when 1% CaCl2 and Pluronic F68 were used. The optimized formulation was coated with chitosan to increase the physical stability of the nanoparticles. The final nanoparticles presented a mean diameter of 321 nm and polydispersity index of 0.217, and spherical shape. After coating, the Zeta potential was +21 mV, and the encapsulation efficiency was 73 %. The in vitro release assay showed that about 98% of the drug was released from the nanoparticles after 48 h. Moreover, the nanoparticles reduced hesperidin cytotoxicity on healthy cells (Vero cells) and improved the cytotoxicity on tumor cells (HeLa, PC-3 and Caco-2 cells). Results showed that the chitosan-coated gliadin nanoparticles are potential carriers for hesperidin delivery for cancer treatment.

Bovine serum albumin-based nanoparticles containing the flavonoid rutin produced by nano spray drying

Rutin is a flavonoid compound obtained from different vegetables and fruits; specifically, it is found in the seeds of buckwheat and in fruit peels, particularly citrus. It is also an important constituent of red wine. Rutin exhibits various biological properties including antiviral, vasoprotective, anti-inflammatory, and anticarcinogenic activities. However, its antioxidant activity is the most well studied. Despite the potential for in vitro applications, rutin presents low oral bioavailability that affects its biological activities. Nanoparticles composed of polymers, protein, or lipids are of great importance in the pharmaceutical and nutraceutical areas due to their physicochemical properties, which improve the pharmacokinetics of the drug which is loaded within. This study presents the production of bovine serum albumin (BSA) nanoparticles containing rutin by nano spray drying. Nanoparticles were characterized in terms of mean particle size, size distribution, morphology, zeta potential, and drug content; as well as their antioxidant activity. The optimized spray-drying conditions produced spherical particles with a mean size of 316 nm, zeta potential of −32 mV, and encapsulation efficiency around 32%. Moreover, when antioxidant activity toward the ABTS+ radical was assayed, nanoencapsulation increased the IC50 of rutin by 2-fold. The nano spray-drying process proved to be suitable for the production of rutin-loaded BSA nanoparticles with potential antioxidant activity.

A stability-indicating HPLC-PDA method for the determination of ferulic acid in chitosan-coated poly(lactide-co-glycolide) nanoparticles

ABSTRACT The development and validation of a simple and efficient method for the quantification of ferulic acid in poly (D,L-lactide-co-glycolide) (PLGA) nanoparticles coated with chitosan (CS) by reverse phase high performance liquid chromatography coupled to photodiode array detection was described. For the chromatographic analysis, a reverse phase C-18 column was used, mobile phase consisting of acetonitrile and 0.5% acetic acid (37:63, v/v), isocratically eluted at a flow rate of 1 mL/min. Drug determination was performed at 320 nm. The method was validated in terms of the selectivity, linearity, precision, accuracy, robustness, limits of detection and quantification. The method was linear in the range of 10 to 100 µg/mL (r=0.999) and presented limit of detection and quantification of 102 ng/mL and 310 ng/mL, respectively. The method was precise (intra and inter-day) based on relative standard deviation values (less than 3.20%). The recovery was between 101.06 and 102.10%. Robustness was demonstrated considering change in mobile phase proportion. Specificity assay showed no interference from the components of nanoparticles or from the degradation products derived from acidic and oxidative conditions. The proposed method was suitable to be applied in determining the encapsulation efficiency of ferulic acid in PLGA-CS nanoparticles and can be employed as stability indicating one.

Aspectos físicos e biológicos de nanopartículas de ferritas magnéticas / Physical and biological aspects of ferrite magnetic nanoparticles

Nanopartículas de óxidos magnéticos são compostas principalmente de Fe3O4 (magnetita) e Fe2O3 (maghemita), Também são muito utilizadas as de CoFe2O4 (ferrita de cobalto), NiFe2O4 (ferrita de níquel), entre outras, As nanopartículas de ferrita apresentam diversas aplicações na área biomédica, entre as quais a liberação controlada de fármacos, agentes de contraste para imagem de ressonância nuclear magnética, transportadores de fármacos guiados por campo magnético, tratamento de tumores via hipertermia, separação biomolecular magnética e diagnóstico, Para que as nanopartículas possam ser utilizadas devem possuir características magnéticas adequadas além de controle no tamanho e composição da superfície, Nesta revisão foi descrito um novo método de síntese e caracterização de nanopartículas de óxidos magnéticos os quais podem ser usados em aplicações biomédicas...

Nanoparticles of magnetic oxides are mainly composed of Fe3O4 (magnetite) and Fe2O3 (maghemite). However, CoFe2O4 (cobalt ferrite) and NiFe2O4 (Niguel ferrite), among others, are highly used. These ferrite nanoparticles show many biomedical applications, among them the controlled drug-release, contrast agents for magnetic resonance imaging, magnetic guided drug carriers to tumors treatment by hyperthermia, magnetic biomolecular separation and diagnostics. For these nanoparticles can be used, it should present adequate magnetic characteristics as well as controlled particle size and surface composition. In this review it was described a new method of synthesis and characterization of nanoparticles, as well as magnetic behavior and biomedical applications of nanoparticles...

Determination of amphotericin B in PLA-PEG blend nanoparticles by HPLC-PDA

In this work, we developed and validated an effective reversed-phase HPLC method with photodiode array (PDA) detection for the quantitative analysis of amphotericin B (AmB) in poly(lactide)-poly(ethylene glycol) (PLA-PEG) blend nanoparticles. Chromatographic runs were performed on a reverse phase C18 column using a mobile phase comprising a 9% acetic acid and acetonitrile mixture (40:60, v/v) under isocratic elution with a flow rate of 1 mL/min. AmB was detected at a wavelength of 408 nm. The validation process was performed considering the selectivity, linearity, precision, accuracy, robustness, limit of detection (LOD) and limit of quantitation (LOQ) of the method. A concentration range of 1-20 µg/mL was used to construct a linear calibration curve. The LOQ and LOD were 55 and 18 ng/mL, respectively. The mean recovery of AmB from the samples was 99.92% (relative standard deviation (RSD) = 0.34%, n=9), and the method was robust for changes in the flow rate of the mobile phase (maximum RSD=4.82%). The intra- and inter-assay coefficients of variation were less than 0.59%. The method was successfully used to determine the entrapment efficiency of AmB in PLA-PEG blend nanoparticles.

Neste trabalho desenvolveu-se e validou-se um efetivo método por cromatografia líquida de alta eficiência (CLAE) em fase reversa com detecção por fotodiodos para a análise quantitativa de anfotericina B (AmB) em nanopartículas compostas por blendas de poli(ácido lático)-polietilenoglicol (PLA-PEG). Corridas cromatográficas foram realizadas sob coluna C18 de fase reversa com fase móvel consistindo de ácido acético 9% e acetonitrila (40:60, v/v), em eluição isocrática com fluxo de 1 mL/min. A AmB foi detectada no comprimento de onda de 408 nm. O processo de validação foi realizado considerando a seletividade, linearidade, precisão, exatidão, robustez, limite de detecção (LD) e limite de quantificação (LQ) do método. Uma faixa de concentração entre 1-20 µg/mL foi usada para obter a curva-padrão linear. Os valores de LD e LQ foram 55 e 18 ng/mL, respectivamente. A recuperação média da AmB a partir das amostras foi de 99,92% (desvio padrão relativo = 0,34%, n=9) e o método foi robusto, considerando alterações no fluxo da fase móvel (desvio padrão relativo máximo=4,82%). Os coeficientes de variação intra e inter dia foram inferiores a 0,59%. O método foi utilizado com sucesso para a determinação da eficiência de encapsulação da AmB em nanopartículas de PLA-PEG.

Efeito da nicotina sobre fagócitos ativados / Effect of the nicotine on actived phagocytes

Indivíduos fumantes apresentam aumento na proporção de leucócitos polimorfonucleares (LPMN), como por exemplo, no tecido pulmonar, resultando em aumento nos níveis de enzimas proteolíticas e espécies reativas de oxigênio, que ocasionam efeito destrutivo na matriz celular e contribuem para a progressão da doença pulmonar, além de favorecer o aparecimento de infecções microbianas, e determinam que as células fagocíticas estejam em frequente estado de ativação (fagocitose). Neste trabalho, avaliou-se a influência da nicotina (NIC) sobre a viabilidade de LPMN e macrófagos ativados ou não, pelos estímulos zymosan e acetato de forbol miristato. Os resultados indicaram que a NIC promove aumento na viabilidade de LPMN e macrófagos ativados em relação a essas células ativadas sem a presença de NIC, avaliada 'ex vivo' pelo teste de exclusão do azul de trypan. Esse efeito foi significativamente mais pronunciado sobre LPMN que sobre os macrófagos. Essa redução na citotoxicidade favorece a sobrevida da célula, podendo exacerbar os seus efeitos deletérios, especialmente em no seu estado ativado, pela maior produção de espécies reativas de oxigênio.

Smokers show increased rates of polymorphonuclear leukocytes (PMNL), including in pulmonary tissue, resulting increased levels of proteolytic enzymes and reactive oxygen species, which have a destructive effect on the cellular matrix and contribute to the progression of pulmonary disease, in addition to promoting the onset of microbial infections which cause phagocytic cells to be in a state of frequent activation (phagocytosis). This work evaluated the influence of nicotine (NIC) on the viability of PMNL and macrophages (activated or not), by stimuli zymosan and phorbol myristate acetate. The results indicated that NIC led to increased viability of PMNL and activated macrophages compared to these cells activated without NIC, measured ex vivo by trypan blue exclusion test. This effect was significantly higher on PMNL than on macrophages. This reduction in cytotoxicity favors cell survival, and may exacerbate its deleterious effect, especially in the active state, due to increased production of reactive oxygen species.

Development and validation of an HPLC method for the determination of fluorouracil in polymeric nanoparticles

The objective of this work was to develop and validate a rapid high performance liquid chromatography (HPLC) method for the quantitative analysis of fluorouracil (5-FU) in polymeric nanoparticles. Chromatographic analyses were performed on an RP C18 column with a mobile phase consisting of acetonitrile and water (10:90, v/v) at a flow rate of 1 mL/min. The 5-FU was detected and quantitated using a photodiode array detector at a wavelength of 265 nm. The method was shown to be specific and linear in the range of 0.1-10 µg/mL (r = 0.9997). The precision (intra- and inter-day) was demonstrated because the maximum relative standard deviation was 3.51%. The method is robust relative to changes in flow rate, column and temperature. The limits of detection and quantitation were 10.86 and 32.78 ng/mL, respectively. The method fulfilled the requirements for reliability and feasibility for application to the quantitative analysis of 5-FU in polymeric nanoparticles.

O objetivo deste trabalho foi desenvolver e validar um método rápido de cromatografia líquida de alta eficiência (CLAE) para análise quantitativa de fluorouracila (5-FU) em nanopartículas poliméricas. Corridas cromatográficas foram realizadas sob uma coluna RP C18 com uma fase móvel consistindo de acetonitrila e água (10:90, v/v) a um fluxo de 1 mL/min. O 5-FU foi detectado e quantificado através de um detector de fotodiodos em um comprimento de onda de 265 nm. O método demonstrou ser específico e linear na faixa de 0,1-10 µg/mL (r =0.9997). As precisões (intra e inter dia) revelaram um desvio padrão relativo máximo de 3,51%. O método é robusto considerando mudanças realizadas no fluxo da fase móvel, temperatura e marca da coluna. Os limites de detecção e quantificação foram de 10,86 e 32,78 ng/mL, respectivamente. O método cumpriu os requisitos para ser considerado confiável e viável para aplicação na análise quantitativa de 5-FU em nanopartículas poliméricas.

Salacia campestris root bark extract: peroxidase inhibition, antioxidant and antiradical profile / Salacia campestris extrato da casca da raiz: inibição da peroxidase, antioxidante e anti-radicalar perfil

Reactive oxygen species (ROS) and free radical species have been implicated in initiating or accompanying many diseases in living organisms; there is thus, a continual need for antioxidants molecules to inactivate ROS/free radicals. Many studies of plants crude extracts have demonstrated free-radical scavenging and antioxidant action. Salacia species have long been used, in several countries, as traditional medicines against certain diseases and for their anti-inflammatory properties. In this study, Salacia campestris Walp (Hippocrateaceae) root bark ethanol extract (ScEtOH) was assessed for its ability to scavenge free radicals and reactive oxygen species; the results were expressed as percentage inhibition of the active species. ScEtOH was efficient against studied species: DPPH radical (obtained inhibition = 30 percent), ABTSò+ (IC50 = 1.8±0.8 μg/mL), HOCl (IC50 = 1.7 ± 0.1 μg/mL), O2ò- (obtained inhibition = 32 percent), and NOò (obtained inhibition = 18 percent). Peroxidase activity inhibition was evaluated through the guaiacol oxidation reaction catalyzed by hemin, HRP and myeloperoxidase (MPO); data showed that ScEtOH at 10 μg/mL led to 54 and 51 percent of inhibition, respectively, for the hemin and HRP systems. In the MPO system, ScEtOH promoted a 50 percent inhibition at 8.9 μg/mL, whereas quercetin, a powerful MPO inhibitor, inhibited this system at 1.35 μg/mL.

Espécies reativas do oxigênio (ERO) e radicais livres estão relacionados ao início ou à exacerbação de muitas doenças em organismos vivos; existindo portanto uma necessidade contínua por moléculas antioxidantes que inativem as ERO e radicais livres. Muitos estudos com extratos brutos de plantas têm demonstrado propriedades antioxidantes e sequestradoras de radicais livres. Espécies de Salacia são utilizadas, em muitos países, como remédio tradicional contra certas doenças e por suas propriedades antiinflamatórias. Neste estudo, o extrato bruto etanólico da casca da raiz da Salacia campestris Walp (Hippocrateaceae) foi avaliado quanto à sua habilidade em seqüestrar radicais livres e espécies reativas do oxigênio; os resultados são expressos como porcentagem de inibição das espécies ativas. ScEtOH mostrou-se eficiente frente as espécies estudadas: radical DPPH (inibição obtida = 30 por cento), ABTSò+ (IC50 = 1,8±0,8 μg/mL), HOCl (IC50 = 1,7 ± 0,1 μg/mL), O2ò- (inibição obtida = 32 por cento), and NOò (inibição obtida = 18 por cento). A inibição da atividade peroxidásica foi avaliada através da oxidação do guaiacol catalisada pela hemina, HRP e mieloperoxidase (MPO); os dados mostram que 10 μg/mL de ScEtOH promovem 54 e 51 por cento de inibição, respectivamente para os sistemas da hemina e da HRP. No sistema da MPO, ScEtOH promoveu 50 por cento de inibição na dose de 8,9 μg/mL, enquanto a quercetina, um potente inibidor da MPO promoveu tal inibição com 1,35 μg/mL.

Profile of Maytenus aquifolium action over free radicals and reactive oxygen species

Reactive oxygen species (ROS) and free radical species have been implicated in initiating, accompanying or causing many diseases in living organisms; there is thus, a continual need for antioxidants molecules to inactivate ROS/free radicals. Many studies of plants crude extracts have demonstrated free-radical scavenging and antioxidant action. Maytenus species have long been used, in several countries, as traditional medicines against gastric ulcers, dyspepsia and others gastric problems and for their anti-inflammatory properties. In this study, Maytenus aquifolium (Celastraceae) root bark ethanol extract was assessed for its ability to scavenge free radicals and reactive oxygen species. The results were expressed as percentage inhibition of the active species. The extract was efficient against studied reactive species: DPPH radical (obtained inhibition = 35.5 ± 1.3 percent), ABTSò+ (IC50 = 0.0036 ± 0.0003 mg/mL), HOCl (IC50 = 0.002 ± 0.0001 mg/mL ), O2ò- (obtained inhibition = 36.0 ± 2.1 percent), and NOò (obtained inhibition = 18.3 ± 0.4 percent).

Espécies Reativas do Oxigênio (ERO) e radicais livres têm tido implicações na iniciação e evolução de muitas doenças ou nas causas das mesmas em organismos vivos; há portanto, necessidade contínua por moléculas antioxidantes para inativar ERO/radicais livres. Estudos sobre extratos brutos de plantas têm demonstrado suas ações antioxidante e seqüestradora de radicais livres. Espécies do gênero Maytenus são utilizadas, em vários países, como medicamentos tradicionais no combate a úlceras gástricas, dispepsia e outras desordens gástricas, bem como por suas propriedades antiinflamatórias. Neste estudo, o extrato bruto etanólico da raiz da Maytenus aquifolium (Celastraceae) foi avaliado quanto à sua habilidade em seqüestrar radicais livres e outras espécies reativas do oxigênio. Os resultados são expressos como porcentagem de inibição das espécies ativas. O extrato foi eficiente contra as espécies estudadas: radical DPPH (inibição alcançada = 35,5 ± 1,3 por cento), ABTSò+ (IC50 = 0,0036 ± 0,0003 mg/mL), HOCl (IC50 = 0,002 ± 0,0001 mg/mL ), O2ò- (inibição alcançada = 36,0 ± 2,1 por cento), and NOÀ (inibição alcançada = 18,3 ± 0,4 por cento).