TNF-α inhibition, antioxidant effects and chemical analysis of extracts and fraction from Brazilian guaraná seed powder.

Paullinia cupana Kunth., commonly named Guaraná, is a plant from Brazil used as stimulant. The aim of this study was to evaluate the potential of extracts and tannins-rich and methylxanthines-free fraction from guaraná in the anti-inflammatory and antioxidant effect in vitro. Extract 1 obtained good yields of tannins and methylxanthines and was used to identify a type-A procyanidin trimer by LC-ESI-MS. Fraction 4 was rich in tannins and absent of methylxanthines. The extracts and fraction exhibited strong capacity for scavenging DPPH radical with IC50 between 5.88 and 42.75-µg/mL and inhibited TNF-α release by LPS-activated THP-1 cells when compared with control cells and did not present toxicity to THP-1 cells. The fraction 4, rich in tannins, was highly active, with IC50 5.88 µg/mL by DPPH method and inhibited TNF-α release in 83.50% at 90 µg/mL. These results reinforced potential anti-inflammatory of guaraná and data for new therapeutic approaches.

A rapid simultaneous determination of methylxanthines and proanthocyanidins in Brazilian guaraná (Paullinia cupana Kunth.).

Paullinia cupana is a plant native to Brazil that is widely used in traditional medicine as a physical and mental stimulant. It is also used worldwide to produce soft drinks. A method for the simultaneous quantitation of seven markers in guaraná by HPLC-PDA was developed, and extraction methods for the determination of methylxanthines and tannins were investigated. Quantified substances were theobromine, theophylline, caffeine, catechin, epicatechin, procyanidins A2 and B2. Results confirmed the satisfactory selectivity and linearity (r2≥0.99) within the mass ranges. Repeatability (RSD≤2.80%), intermediate precision (RSD≤4.47%), accuracy (recoveries from 90.59%-104.67%), and robustness were demonstrated. Extract 1 presented the contents: 0.0177% (±1.02%) for theobromine, 0.0131% (±1.14%) for theophylline, 2.9429% (±1.27%) for caffeine, 0.4563% (±1.02%) for catechin, 0.5515% (±1.05%) for epicatechin, 0.0607% (±2.80%) for A2 and 0.1035% (±1.39%) for B2. The method for simultaneous quantitation of seven chemical markers in guaraná proved to be reliable using a simple and convenient HPLC setup.

Development and validation of alternative methods by non-aqueous acid-base titration and derivative ultraviolet spectrophotometry for quantification of sildenafil in raw material and tablets

Sildenafil citrate (SILC) is a potent phosphodiesterase-5 inhibitor used for erectile dysfunction and pulmonary hypertension. This study shows two simple, fast and alternative analytical methods for SILC determination by non-aqueous titration and by derivative ultraviolet spectrophotometry (DUS) in active pharmaceutical ingredient and/or dosage forms. The quantitation method of SILC active pharmaceutical ingredient by non-aqueous acid-base titration was developed using methanol as solvent and 0.1 mol/L of perchloric acid in acetic acid as titrant. The endpoint was potentiometrically detected. The non-aqueous titration method shows satisfactory repeatability and intermediate precision (RSD 0.70-1.09%). The neutralization reaction occurred in the stoichiometric ratio 1:1 in methanol. The determination of SILC active pharmaceutical ingredient or dosage forms by DUS was developed in the linear range from 10 to 40 µg/mL, in 0.01 mol/L HCl, using the first order zero-peak method at λ 256 nm. The DUS method shows selectivity toward tablets excipients, appropriate linearity (R2 0.9996), trueness (recovery range 98.86-99.30%), repeatability and intermediate precision in three concentration levels (RSD 1.17-1.28%; 1.29-1.71%, respectively). Therefore, the methods developed are excellent alternatives to sophisticated instrumental methods and can be easily applied in any pharmaceutical laboratory routine due to simple and fast executions.

Stability-indicating HPLC-UV for the determination of 4-bromomethyl-3-nitrobenzoic acid, a bioactive nitrocompound.

The nitroaromatic compound 4-bromomethyl-3-nitrobenzoic acid (ANB) is a promising antitumoral agent whose activity has recently been investigated. Forced degradation studies were conducted on ANB with a high-performance liquid chromatography-ultraviolet assay to establish its stability and selectivity. ANB was subjected to degradation studies under hydrolytic (acid and alkaline), oxidative and light exposition conditions. The compound showed greater lability only in acid and alkaline conditions by forming a major degradation product. The chromatographic separation of ANB and its degradation product was achieved on an octadecylsilane column using a mobile phase of methanol-water (80:20, v/v), pH 4.0 adjusted with formic acid, flow rate of 0.7 mL/min, ultraviolet diode array detection at λ 271 nm, injection volume of 20 µL and temperature of 30°C. The method was validated according to International Conference on Harmonization guidelines with respect to precision (average relative standard deviation 0.67%), accuracy (average 99.97%), linearity (y = 118730x + 12912; coefficient of determination > 0.999), specificity and robustness (p > 0.05). The degradation product formed as a result from the hydrolysis of nitrobenzyl bromide, corresponded to its benzyl alcohol, 4-hydroxymethyl-3-nitrobenzoic acid (ANOH) and was characterized by co-elution with a synthetic ANOH working standard.

Evaluation of antitumor activity and development of solid lipid nanoparticles of metronidazole analogue.

Nitroheterocyclic compounds have received considerable interest as hypoxia-selective cytotoxins (HSC) for cancer treatment. In the present study, we investigated antitumor activity of an iodide analogue of metronidazole, 1-(2-iodoethyl)-2-methyl-5-nitroimidazole (MTZ-I), using Swiss mice bearing solid Ehrlich tumor. MTZ-I showed potent anti-cancer activity at a dose of 40 mg/kg. MTZ-I loaded solid lipid nanoparticles (SLN) were developed as an alternative colloidal carrier system to enhance tumor drug uptake. SLN were characterized for particle size, polydispersity index, zeta potential and entrapment efficiency. In addition, the influence of presence of the cationic lipid stearylamine (STE) on stability of formulation was assessed. The results of DSC study showed that MTZ-I exhibited interaction with STE.

A qualidade, o custo e a dispensação de sinvastatina cápsulas em farmácias magistrais / Quality, cost and dispensing of simvastatin capsules at compounding pharmacies

Com base na RDC/ANVISA 67/2007, avaliou-se o serviço prestado na dispensação de sinvastatina cápsulas em farmácias magistrais. Vinte e quatro (16%) das 150 farmácias magistrais foram aleatoriamente selecionadas nas regiões de Belo Horizonte-MG, observando a distribuição geográfica. A qualidade do produto, os aspectos da dispensação e o custo foram considerados em estudos de correlação. Os resultados foram submetidos a análises estatísticas descritivas e inferenciais. Em apenas 25% destas farmácias, o fax da prescrição foi solicitado. Todos os pedidos foram entregues no prazo, porém sem comprovação da dispensação. Em uma das entregas, o medicamento era o incorreto. Seis (25%) dos produtos apresentaram rotulagem incompleta (ausência de posologia 16,3% ou de identificação do médico/paciente 8,7%). Apenas 11 (45,8%) das amostras apresentaram qualidade satisfatória. O custo do medicamento manipulado variou entre R$20,40 e R$44,00 com valor médio R$31,18. Não se observou associação entre a qualidade do medicamento e os aspectos do custo, prazo de entrega e localização geográfica. Verifica-se que a maioria das farmácias magistrais ainda precisa adequar-se à legislação para garantir à população uma assistência farmacêutica de qualidade.

The aim of the present study was to evaluate the dispensing of simvastatin capsules at compounding pharmacies in light of Brazilian public health regulations (RDC/ANVISA 67/2007). Twenty-four (16%) out of 150 compounding pharmacies were randomly selected in the city of Belo Horizonte based on geographic distribution among the different districts. Dispensing services, quality and cost were considered in the correlation studies. The results were submitted to descriptive and inferential statistical analyses. The prescription fax was requested in only 25% of the pharmacies. All orders were delivered on time, but without proof of dispensing. The medication was incorrect in one of the deliveries. Six (25%) product labels were incomplete (16.3% for absence of drug regimen; 8.7% for absence of physician/patient identification). Only 11 (45.8%) of the samples exhibited satisfactory quality. The cost of the compounded drug ranged from R$20.40 to R$44.00 (mean: R$31.18). No association was found between drug quality and cost, delivery or geographic location. In conclusion, the majority of compounding pharmacies need to make adjustments in order to meet the requirements of Brazilian legislation and ensure the quality of pharmaceutical services offered to the population.

Stryphnodendron adstringens (Mart. ) Coville (Fabaceae) proanthocyanidins quantitation by RP-HPLC

Stryphnodendron adstringens (Mart.) Coville (barbatimão) is a tree belonging to the Fabaceae family, and it is commonly found in the southeastern Brazilian cerrado. The stem bark of this tree is traditionally used as an anti-inflammatory and antiseptic agent to treat leucorrhea and diarrhea, as well as to promote wound healing (owing to the presence of proanthocyanidins). Proanthocyanidins were obtained from the ethanolic extract of S. adstringens stem bark and assessed by reversed phase-high performance liquid chromatography with an ultraviolet/diode array detector. The identified compounds included gallic acid, catechin, gallocatechin (GC), epigallocatechin, and epigallocatechin gallate (EGCG). The selected markers, GC and EGCG, were simultaneously used for chromatographic validation (linearity range: 30-330 ng, equivalent to 3-33 µg/mL; r>0.998). The method showed precision (intra-day relative standard deviation [RSD]: 1.72% for GC; 1.16% for EGCG; inter-day RSD: 1.74%-2.60% for both markers), accuracy, robustness, and selectivity. The limits of detection and quantitation were 0.29 µg/ml and 0.89 µg/ml for GC, and 0.88 µg/mL and 2.67 µg/mL for EGCG, respectively. In addition, S. obovatum was evaluated and showed an average of 12.2 µg/mL for GC (equivalent to 1.22% w/w) and 14.2 µg/mL for EGCG (equivalent to 1.42% w/w) in the ethanolic extract. The quantitative results were compared to those obtained for S. adstringens, which showed that the markers are present in both species.

Stryphnodendron adstringens (Mart.) Coville (barbatimão) é uma árvore da família Fabaceae encontrada no cerrado do sudeste do Brasil. As cascas do caule das espécies são tradicionalmente utilizadas como agente antiinflamatório e antisséptico para o tratamento de diarreia e leucorreia, bem como para promover a cicatrização de feridas, devido à presença de proantocianidinas. Estas substâncias obtidas a partir do extracto etanólico (EE) de cascas do caule de S. adstringens foram avaliadas por cromatografia líquida de alta eficiência de fase reversa. As substâncias identificadas foram ácido gálico, catequina, galocatequina (GC), epigalocatequina e galato de epigalocatequina (EGCG). Os marcadores selecionados, GC e EGCG, foram utilizados simultaneamente para validação cromatográfica (intervalo de linearidade 30-330 ng, equivalente a 3-33 µg/mL, r>0,998). O método mostrou precisão (intra-dia DPR 1,72% para GC; 1,16% para EGCG; inter-dia DPR 1,74%-2,60% para ambos os marcadores), exatidão, robustez e seletividade. Os limites de detecção e de quantificação de 0,29 µg/mL e 0,89 µg/mL para a GC, e 0,88 ng/mL e 2,67 µg/mL para EGCG, respectivamente. Outra espécie (S. obovatum) avaliada, mostrou valores médios 12,2 µg/mL de GC (equivalente a 1,22% p/p) e 14,2 µg/mL de EGCG (equivalente a 1,42% p/p) encontrados no EE. Os resultados quantitativos comparam-se aos de S. adstringens e mostram que tais marcadores estão presentes em ambas as espécies.

Quality evaluation of simvastatin compounded capsules / Avaliação da qualidade de cápsulas manipuladas de sinvastatina

Simvastatin is commercially available as tablets and compounded capsules in Brazil. Very few reports regarding these capsules' quality, and consequently their efficacy, are available. The pharmaceutical quality of 30 batches of 20 mg simvastatin capsules from the market was evaluated by weight determination, content uniformity, disintegration (Brazilian Pharmacopeia), assay and dissolution test (USP32 tablet monograph). A HPLC method was developed for assay, content uniformity and dissolution test, and specifications were also established. Out of the 30 batches evaluated, 29 showed capsule disintegration within 45 min and individual weight variation was within ±10 percent or ±7.5 percent relative to average weight, for < or > 300 mg, respectively. Only 27 batches met dissolution test criteria with values >80 percent of the labeled amount in 45 min; 21 batches showed simvastatin content between 90.0-110.0 percent of the labeled amount and 19 batches had at least 9 out of 10 capsules with content uniformity values between 85.0-115.0 percent of the labeled amount with RSD<6.0 percent. Only 14 of all (30) batches fully met pharmacopeial quality standards. The establishment of test conditions and specification parameters for simvastatin capsules showed that there are relevant pharmacopeial quality differences between batches compounded by different pharmacies. For 53.33 percent of the tested products hypercholesterolemic treatment efficacy may be compromised.

No Brasil, a sinvastatina está comercialmente disponível na forma de comprimidos e cápsulas manipuladas. Poucos relatos estão disponíveis sobre a qualidade e, consequentemente, a eficácia dessas cápsulas. A qualidade de 30 lotes de sinvastatina 20 mg cápsulas do mercado foi avaliada através da determinação de peso, uniformidade de conteúdo, desintegração (Farmacopéia Brasileira), doseamento e teste de dissolução (monografia comprimidos USP32). Método por CLAE foi desenvolvido para o doseamento, uniformidade de conteúdo e teste de dissolução; além disso, especificações foram estabelecidas. Dos 30 lotes avaliados, 29 apresentaram desintegração da cápsula até 45 min e a variação do peso individual foi ± 10 por cento ou ± 7,5 por cento em relação ao peso médio, se < ou > 300 mg, respectivamente. Apenas 27 lotes preencheram os critérios do teste de dissolução com valores > 80 por cento da quantidade rotulada, em 45 min, 21 lotes apresentaram conteúdo de sinvastatina entre 90,0-110,0 por cento do valor rotulado e 19 lotes apresentaram pelo menos 9 em 10 cápsulas, com valores de uniformidade de conteúdo entre 85,0 -115,0 por cento da quantidade rotulada com RSD < 6,0 por cento. Apenas 14 de todos os lotes (30) atenderam completamente os padrões de qualidade farmacopéicos. O estabelecimento das condições para os testes e de especificações para os parâmetros das cápsulas de sinvastatina mostrou que houve diferenças relevantes na qualidade farmacopeica entre os lotes das cápsulas manipuladas por distintas farmácias. A eficácia do tratamento hipercolesterolêmico poderia estar comprometida para 53,33 por cento dos produtos testados.

Caracterização do fármaco hipoglicemiante glibenclamida / Characterization of the hypoglycemic drug glibenclamide

A glibenclamida (GLIB) é um fármaco de segunda geração, administrado por via oral na forma de comprimidos, utilizado para o tratamento de Diabetes mellitus. GLIB possui baixa solubilidade aquosa, o que pode levar a uma baixa liberação a partir de formas farmacêuticas sólidas no teste de dissolução e, portanto, a variabilidades no tratamento. Neste estudo, avaliam-se as características da matéria-prima GLIB, que podem influenciar o perfil de dissolução, e conseqüentemente, a biodisponibilidade, por meio de técnicas tais como, adsorção de nitrogênio, difração de raio laser, análise térmica, espectroscopia por IV/UV e difração de raios X.

Glibenclamide (GLIB) or glyburide, a second-generation hypoglycemic agent is orally used in the form of tablets for the treatment of diabetes mellitus. Bulk GLIB has a low aqueous solubility and it may yield low drug release in the dissolution test, causing variabilility in the treatment. This work evaluates the bulk GLIB features, which may influence drug release profile, hence, bioavailability, by means of techniques such as nitrogen sorption analysis, laser diffraction, thermal analysis, IV/UV spectroscopy and X-ray analysis.

Determinação de daidzeína, genisteína e gliciteína em cápsulas de isoflavonas por cromatografia em camada delgada (CCD) e cromatografia líquida de alta eficiência (CLAE) / Determination of daidzein, genistein and glycitein in isoflavone capsules by thin layer chromatography (TLC) and high performance liquid chromatography (HPLC)

A utilização de produtos naturais e suplementos contendo extratos secos de isoflavonas aumentou na última década, devido, principalmente, aos efeitos benéficos atribuídos a estes compostos no alívio dos sintomas da menopausa. Genisteína, daidzeína e gliciteína são as agliconas mais abundantes nos extratos de soja, ocorrendo também como glicosídeos. Tendo em vista seus usos, não existe ainda na literatura uma definição da quantidade mínima de cada uma das agliconas que os extratos ou cápsulas de isoflavonas devem ter, e também não existe um método oficial para o controle de qualidade dos mesmos. O presente trabalho apresenta um método por CCD para análise qualitativa das três agliconas e de seus glicosídeos em extratos e cápsulas de isoflavonas, antes e após hidrólise ácida. A análise quantitativa das cápsulas de isoflavonas, realizada por cromatografia líquida de alta eficiência (CLAE), revelou grandes variações nos teores das três agliconas, após hidrólise ácida. Os teores variaram da seguinte forma, nos 18 lotes de cápsulas avaliados: daidzeína (13,34 a 76,20 mg/cápsula), genisteína (0,61 a 27,18 mg/cápsula) e gliciteína (0,49 a 8,80 mg/cápsula).

The use of herbal products and supplements based on isoflavone dry extracts has increased considerably in the last decade, mainly due to beneficial effects to relief of the menopausal symptoms credited to those compounds. Genistein, daidzein and glycitein are the most abundant isoflavone aglycones found in soy extract, where they also occur as glycosides. Concerning their uses, there is neither standardization regarding the minimum content of each aglycone in the extracts or capsules, nor an official method to the quality control of these products. The present work presents a TLC method for qualitative analysis of the three aglycones and their glycosides in extracts and capsules of isoflavones, before and after acid hydrolysis. The quantitative analysis of the isoflavone capsules, carried out by high performance liquid chromatography (HPLC), showed high variation in the content of the three aglycones, after acid hydrolysis. The contents varied in the following way, in the 18 batches of evaluated capsules: daidzein (13.34 to 76.20 mg/capsule), genistein (0.61 to 27.18 mg/capsule) and glycitein (0.49 to 8.80 mg/capsule).

Teste de dissolução para avaliação de liberação de glibenclamida em comprimidos / Glibenclamide dissolution test for drug release evaluation in tablets

A glibenclamida (GLIB) ou gliburida, é um hipoglicemiante oral de segunda geração, da classe das sulfoniluréias, usado sob a forma de comprimidos para tratamento do diabetes mellitus. Variações no tratamento podem ocorrer, devido à baixa solubilidade do fármaco em comprimidos. A comparação de várias formulações de comprimidos piloto com comprimidos do medicamento referência (Daonil®, glibenclamida 5 mg comprimidos, Aventis Pharma Ltda.) foi avaliada por meio do desenvolvimento de um teste de dissolução sem adição de solventes orgânicos ou tensoativos no meio, que mostrou ser discriminativo para as diferentes formulações farmacêuticas propostas. A quantificação de GLIB foi realizada por meio de cromatografia líquida de alta eficiência em fase reversa (CLAE-FR), método previamente validado. A partir de vários ensaios de perfil de dissolução testados comparativamente àquele de comprimidos do medicamento referência, verificou-se o potencial de determinada formulação proposta (f1 4,04 and f2 69,35) como candidata a medicamento genérico no mercado brasileiro.

Glibenclamide (GLIB) or glyburide, a second-generation hypoglycemic agent is used per oral as tablets for the treatment of diabetes mellitus. Much variabilility in the treatment may occur because of the low drug aqueous solubility in tablets dosage forms. This work reports the comparison of several pilot formulation batches with the commercial reference drug dosage form (Daonil®, glibenclamide 5 mg per tablet, Aventis Pharma Ltda.). A feasible dissolution test, developed with no use of organic solvents or surfactants in the medium, showed to be discriminative regarding to different formulations tested. GLIB quantitation was performed by a previously validated reverse-phase high performance liquid chromatography (RP-HPLC). Among several dissolution profiles compared with that of a commercial reference, a potential for a generic candidate was evident (f1 4.04 and f2 69.35) for a proposed solid dosage formulation in the Brazilian market.

Determinação da glibenclamida por espectrofotometria derivada no ultravioleta para avaliação do teste ou perfil de dissolução de comprimidos / Glibenclamide determination by derivative ultraviolet spectrophotometry for test or dissolution profile assessment in tablets

A glibenclamida (GLIB) ou gliburida, agente hipoglicemiante oral de segunda geração, da classe das sulfoniluréias, é usada sob a forma de comprimidos para controle do diabetes mellitus. Possui baixa solubilidade aquosa, podendo proporcionar baixa liberação no teste de dissolução de comprimidos e acarretar variabilidades no tratamento. A solução metanólica de GLIB apresenta máximos de absorção em lambda 210 nm, 227,5 nm e 300 nm na região ultravioleta (UV). Após liberaçao de comprimidos, a sua determinação espectrofotométrica na região UV é dificultada devido à baixa concentração proporcionada após o teste de dissolução (comprimidos GLIB 5 mg, volume de meio 900 mL, 0,56 mg por cento) e à baixa absortividade em comprimento de onda característico, lambda 300 nm. Por estas razões, até hoje, não existe um método de análise proposto para o teste de dissolução em farmacopéias nacional ou estrangeiras para monografia de glibenclamida comprimidos. A falta de uniformidade nestes procedimentos onera a execução de testes de dissolução para verificação da equivalência farmacêutica de especialidades candidatas a genéricos. Este trabalho propõe a determinação espectrofotométrica de GLIB por derivada de primeira e segunda ordens (máximos lambda 238 nm e 218 nm, respectivamente, em tampão fosfato 0,1 mol L-1), utilizando comprimidos referência e teste contendo 5 mg de fármaco por dose unitária.

Glibenclamide (GLIB) or glyburide, a second-generation oral hypoglycemic drug, is used in the tablet form for the treatment of diabetes mellitus. Because of the low aqueous solubility of this sulfonylurea, a low drug release in the dissolution test may occur, hence, causing variabilility in the treatment. GLIB methanolic solution shows UV absorption maxima in lambda 210 nm, 227.5 nm and 300 nm. After released from tablets in the dissolution test, its spectrophotometric determination is difficult due to the low drug concentration in solution (5 mg GLIB tablets, medium volume 900 mL, 0.56 mg percent). In addition, it presents a low absorption in the characteristic wavelength of lambda 300 nm. Up to date, there is no method recommendation for GLIB determination in tablet monographs in many of national or foreign official compendia. For this reason, the dissolution test performance lacks uniformity by official recommended procedures, to prove pharmaceutical equivalence to the reference drug, for generic candidates. GLIB spectrophotometric determination is investigated by first and second derivative orders (peaks at lambda 238 nm and 218 nm, respectively in 0.1 mol L-1 phosphate buffer solution) using reference and test tablets containing 5 mg of GLIB per unit dose.

Saccharin analysis in pharmaceutical and cosmetic preparations by derivative ultraviolet spectrophotometry

Saccharin, a synthetic sweetener constituent of several pharmaceutical and cosmetic preparations, is analyzed qualitatively and quantitatively in the presence of interferences like active ingredients and excipients. A derivative ultraviolet spectrophotometric method employing the second and the fourth orders was used. The calibration data obtained were satisfactory, with a relative precision range of 0.53 por cento-0,72 por cento, and linearity between 0.9998-0.9999.The recovery test results for the toothgel sample ranged between 97.80 por cento and 104.95 por cento in the second order of derivative and between 98.04 por cento and 101.32 por cento in the fourth derivative...

Derivative ultraviolet spectrophotometry: an alternative method for determination of lidocaine hydrochloride in pharmaceutical preparations

Um método alternativo por espectrofotometria derivada de segunda e terceira ordens é descrito, com o objetivo de determinar o cloridrato de lidocaína em produtos não comercializados. Empregaram-se os métodos zero-pico (ZP) e pico-pico (PP) para fins de padronização e quantificação em controle analítico de qualidade. As curvas de calibração obtidas apresentaram linearidade e coeficiente de correlação 0,9999, no intervalo de concentração de 150,0-500,0 µg `mL POT. -1ï. Os desvios padrão relativos médios entre os métodos ZP e PP para a segunda e terceira ordens de derivada foram 0,457 por cento e 0,322 por cento respectivamente. Na aplicação do método à amostra solução injetável, as médias dos coeficientes de variação para a segunda e terceira ordens de derivada foram 0,456 por cento e 0,339 por cento, respectivamente...