Illumina sequencing and assessment of new cost-efficient protocol for metagenomic-DNA extraction from environmental water samples

Abstract In this study, the development and assessment of a modified, efficient, and cost-efficient protocol for mDNA (metagenomic DNA) extraction from contaminated water samples was attempted. The efficiency of the developed protocol was investigated in comparison to a well-established commercial kit (Epicentre, Metagenomic DNA Isolation Kit for Water). The comparison was in terms of degree of shearing, yield, purity, duration, suitability for polymerase chain reaction and next-generation sequencing in addition to the quality of next-generation sequencing data. The DNA yield obtained from the developed protocol was 2.6 folds higher than that of the commercial kit. No significant difference in the alpha (Observed species, Chao1, Simpson and PD whole tree) and beta diversity was found between the DNA samples extracted by the commercial kit and the developed protocol. The number of high-quality sequences of the samples extracted by the developed method was 20% higher than those obtained by the samples processed by the kit. The developed economic protocol successfully yielded high-quality pure mDNA compatible with complex molecular applications. Thus we propose the developed protocol as a gold standard for future metagenomic studies investigating a large number of samples.

Limiar de positividade e sensibilidade dos métodos de Faust et al. e Lutz para detecção de cistos de Giardia duodenalis / Positivity threshold and sensitivity of methods of Faust et al. and Lutz for detection of Giardia duodenalis cysts

Objetivo: O presente estudo teve como objetivo determinar e comparar o limiar de positividade e a sensibilidade dos métodos de centrífugo-flutuação em sulfato de zinco (Faust et al.) e sedimentação espontânea (Lutz) para o diagnóstico de cistos de Giardia duodenalis. Métodos: Para obtenção de amostras fecais com quantidades conhecidas de cistos de G. duodenalis, amostras positivas para o parasito foram purificadas e quantificadas, e posteriormente alíquotas com diferentes quantidades foram adicionadas a amostras fecais negativas para parasitos. Após a contaminação de oito amostras negativas com quantidades variando entre 1.000 e 200.000 cistos por grama de fezes (c/g/f), as mesmas foram submetidas aos métodos de Faust et al. e Lutz, onde o primeiro se mostrou mais sensível para a detecção de cistos de G. duodenalis. Resultados: O limiar de positividade do método de Faust et al. foi de 11.000 c/g/f, e do método de Lutz foi de 100.000 c/g/f, portanto, cargas parasitárias inferiores a esses limiares levariam a resultados falso-negativos. Conclusão: O método de Lutz não é adequado para o diagnóstico de giardíase, portanto deve ser sempre utilizado em conjunto com o método de Faust et al.

Objective: The present study aimed to determine and compare the positivity threshold and sensitivity of the methods of zinc sulfate centrifugal flotation (Faust et al.) and spontaneous sedimentation (Lutz) for the diagnosis of Giardia duodenalis. Methods: To obtain fecal samples containing known amounts of G. duodenalis cysts, the samples with the parasite were purified and quantified, then aliquots with different amounts were added to fecal samples negative for parasites. After the contamination of eight negative samples with amounts ranging between 1.000 and 200.000 cysts per gram of feces, they were subjected to methods of Faust et al. and Lutz, where the first was more sensitive for the detection of G. duodenalis cysts. Results: The positivity threshold of the method of Faust et al. was 11.000 c/g/f, and the method of Lutz was 100.000 c/g/f, so parasitic loads below those thresholds would lead to false-negative results. Conclusion: The method of Lutz is not suitable for the diagnosis of giardiasis, therefore must always be associated with the method Faust et al.

Determinación de Levetiracetam en suero humano por cromatografía líquida con detección de arreglo de diodos / Levetiracetam determination in human serum by liquid chromatography with diode array detection / Determinação de Levetiracetam em soro humano por cromatografia líquida com detecção de arranjo de diodos

Se desarrolló y validó un nuevo método analítico para determinar Levetiracetam (LEV) en suero humano utilizando cromatografía líquida de alta resolución (HPLC) con detección de arreglo de diodos. El procedimiento es sencillo, puede ser incluido en la rutina del laboratorio y prestar servicio tanto en el monitoreo terapéutico como en la urgencia. El método incluye las siguientes etapas: extracción líquido-líquido con diclorometano y evaporación de la fase orgánica, la droga se reconstituye con fase móvil, se inyecta en el cromatógrafo y se detecta a 205 nm. El tiempo de retención de LEV es de 5 minutos y no presenta interferentes con respecto a otras drogas comúnmente prescriptas con Levetiracetam. La curva de trabajo presentó un rango de linealidad entre 5,2 y 82,9 μg/mL, un límite de detección y cuantificación de 0,8 μg/mL y 2,7 μg/mL, respectivamente. La recuperación fue del 99,8%.

A new analytical method for Levetiracetam (LEV) determination in human serum was developed and validated by high performance liquid chromatography (HPLC) with diode detection. It is a simple methodology that can be included in the laboratory routine and can be useful in both therapeutic drug monitoring and emergencies. The drug extraction is performed through a liquid-liquid extraction with methyl chloride. Subsequently, the organic phase is evaporated, reconstituted with the mobile phase, and injected in the chromatograph to be detected at 205 nm. LEV retention time is 5 min and it does not show interference with respect to other drugs commonly prescribed with Levetiracetam. The work curve showed linearity between 5.2 and 82.9 μg/mL and a detection and quantification limit of 0.8 μg/mL and 2.7 μg/mL, respectively, while the recovery was of 99.8%.

Foi desenvolvido e validado um novo método analítico para determinar Levetiracetam (LEV) em soro humano, utilizando cromatografia líquida de alta eficiência (HPLC) com detecção de arranjo de diodos. O procedimento é simples, pode ser incluído na rotina do laboratório e prestar serviço tanto na monitorização terapêutica quanto na urgência. O método inclui as seguintes etapas: extração líquido-líquido com diclorometano, e evaporação da fase orgânica, o fármaco é reconstituído com fase móvel, é injetado no cromatógrafo e detectado a 205 nm. O tempo de retenção de LEV é de 5 minutos e não apresenta interferentes com relação a outras drogas, comumente prescritas com Levetiracetam. A curva de trabalho apresentou um intervalo de linearidade entre 5.2 a 82.9 μg/mL, um limite de detecção e quantificação de 0.8 μg/mL e 2.7 μg/mL respectivamente. A recuperação foi de 99.8%.

Development and validation of an analytical method using High Performance Liquid Chromatography (HPLC) to determine ethyl butylacetylaminopropionate in topical repellent formulations

ABSTRACT Diseases caused by insects are frequent in poor countries, leading to epidemic scenarios in urban areas; e.g., Dengue, Zika and Chikungunya. For this reason, the development of a safe and efficient topical formulation is essential. Ethyl butylacetylaminopropionate (EB) is a mosquito repellent developed by Merck, which is used in products for adults, children and especially babies, due to its low allergenic potential. The aim of this work was to validate an analytical methodology to quantify EB in a new poloxamer-based formulation by high-performance liquid chromatography (HPLC). The quantification methodology was performed at 40 ºC using a Kromasil reverse-phase column (C18), with the dimensions of 250 x 4.6 mm. The mobile phase was acetonitrile:water (1:1) at a 1.0 mL/min flow-rate. The detector wavelength was set at 218 nm to detect EB. The methodology was considered validated since the results indicated linearity (R2>0.99), specificity, selectivity, precision and accuracy (active recovery between 98% and 102%). It also presented limits of detection and quantification of 0.255 µg/mL and 0.849 µg/mL, respectively. The present study demonstrated the EB vehiculated in poloxamer gel is promising as a new insect repellent formulation, since it could be quantified and quality control evaluated.

Assessment of blood sample stability for complete blood count using the Sysmex XN-9000 and Mindray BC-6800 analyzers

Background: Different hematological analyzers have different analytical performances that are often reflected in the criteria for sample stability of the complete blood count. This study aimed to assess the stability of several hematological parameters using the XN-9000 Sysmex and BC-6800 Mindray analyzers. Methods: The impact of storage at room temperature and 4 ◦C was evaluated after 2, 4, 6, 8, 24, 36 and 48h using ten normal and 40 abnormal blood samples. The variation from the baseline measurement was evaluated by the Steel­Dwass­Critchlow­Fligner test and by Bland­Altman plots, using quality specifications and critical difference as the total allowable variation. Results: Red blood cells and reticulocyte parameters (i.e. hematocrit, mean corpuscular volume, mean corpuscular hemoglobin concentration, red blood cell distribution width, immature reticulocyte fractions, low-fluorescence reticulocytes, middle-fluorescence reticulocytes, high fluorescence mononuclear cells) showed less stability compared to leukocyte and platelet parameters (except for monocyte count and mean platelet volume). The bias for hematocrit, mean corpuscular volume, mean corpuscular hemoglobin concentration and red blood cell distribution width coefficient of variation was higher than the critical difference after 8h using both analyzers. Conclusion: Blood samples measured with both analyzers do not show analytically signifi- cant changes in up to 2h of storage at room temperature and 4 ◦C. However, the maximum time for analysis can be extended for up to 8h when the bias is compared to the critical difference

Detection of papillomavirus DNA in formalin-fixed paraffin-embedded equine aural plaque samples / Detecção do DNA de papilomavírus em amostras de placa aural equina fixadas em formalina e embebidas em parafina

A placa aural é uma dermatopatia associada à quatro Equus caballus papillomavirus (EcPVs). Até o momento, o DNA de EcPVs não foi identificado em amostras de placa aural fixadas em formalina e embebidas em parafina (FFPE). O objetivo deste estudo foi otimizar um método para a detecção dos quatro tipos de EcPVs em 21 amostras FFPE usando a PCR. O DNA dos EcPVs foram detectados em 11 amostras (52.4%). O DNA do EcPV4 foi detectado em 38.1% (8/21) e do EcPV3 em 4.8% (1/21) das amostras. Coinfecção foi identificada em duas amostras (9.5%); EcPV4 e 5 foram detectados simultaneamente em uma amostra, enquanto o DNA dos EcPV4 e 6 foi detectado em outra. A especificidade do DNA dos papilomavírus equinos foi avaliada por sequenciamento gênico direto, que confirmou a especificidade dos produtos. A metodologia de PCR proposta possibilita o diagnóstico dos EcPV3, 4, 5 e 6 em amostras FFPE de placa aural equina.(AU)

Effects of different extracts of the mushroom Agaricus blazei Murill on the hematologic profile of mice with Ehrlich tumor / Efeitos de diferentes extratos do cogumelo Agaricus blazei Murill sobre o perfil hematológico de camundongos com tumor de Ehrlich

The aim of this study was to investigate the effect of the mushroom Agaricus blazeii Murril (ABM) extracts on the hematological profile of Swiss mice bearing an Ehrlich solid tumor. Three fractions (total extract, polysaccharides, and supernatant) of ABM extracts obtained by four methods (ultrasonic or water bath, at pH 4 or pH 7) were administered to mice over 21 days. Polysaccharide solutions were analyzed by gas and liquid chromatography that showed both mannose and glucose concentrations. The method of extraction influenced the degree of glucose polymerization and the mannose/glucose relationship. The treatment with ABM supernatant at pH 7 and water bath was associated with reduced concentrations of leukocytes and lymphocytes and altered the percentage of CD4+ and CD8+ lymphocytes in Ehrlich tumor-bearing mice. The treatment with the ABM extract in water bath and ultrasound at pH 4 resulted in lower lymphocyte counts, regardless of tumor presence, and greater granulocyte values in mice with Ehrlich tumor than in controls. We concluded that different fractions and methods of extraction of A. blazei produced differing blood profiles in mice inoculated with the Ehrlich tumor.

O objetivo deste estudo foi investigar o efeito de diferentes extratos do cogumelo Agaricus blazeii Murril (ABM) sobre o perfil hematológico de camundongos Swiss portadores de tumor de Ehrlich sólido. Três frações (extrato total, polissacarídeos e sobrenadante) dos extratos de ABM foram obtidas por quatro métodos (sonificador, banho-maria, em pH 4 ou pH 7) e administradas para camundongos durante 21 dias. Soluções de polissacarídeos foram analisadas por cromatografia gasosa e líquida, que mostraram concentrações de glucose e manose. O método de extração influenciou o grau de polimerização da glicose e a relação manose/glucose. O tratamento com o sobrenadante de ABM (em pH 7 e banho-maria) estava associado com reduzidas concentrações de leucócitos e linfócitos, além de alterar a porcentagem de linfócitos CD4+ e CD8+ em camundongos portadores de tumor sólido de Ehrlich. O tratamento com extratos de ABM, obtidos tanto em banho-maria como no sonificador em pH 4, resultou nas mais baixas contagens de linfócitos, independentemente da presença do tumor, e nos maiores valores de granulócitos em camundongos com tumor de Ehrlich. Conclui-se que os diferentes métodos de extração com as respectivas frações de A. blazei são capazes de intereferir no perfil hematológico de camundongos com tumor sólido de Ehrlich.

Effects of ultrahigh pressure extraction on yield and antioxidant activity of chlorogenic acid and cynaroside extracted from flower buds of Lonicera japonica / 中国天然药物

The present study was designed to establish and optimize a new method for extracting chlorogenic acid and cynaroside from Lonicera japonica Thunb. through orthogonal experimental designl. A new ultrahigh pressure extraction (UPE) technology was applied to extract chlorogenic acid and cynaroside from L. japonica. The influential factors, including solvent type, ethanol concentration, extraction pressure, time, and temperature, and the solid/liquid ratio, have been studied to optimize the extraction process. The optimal conditions for the UPE were developed by quantitative analysis of the extraction products by HPLC-DAD in comparison with standard samples. In addition, the microstructures of the medicinal materials before and after extraction were studied by scanning electron microscopy (SEM). Furthermore, the extraction efficiency of different extraction methods and the 2, 2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activities of the extracts were investigated. The optimal conditions for extracting chlorogenic acid and cynaroside were as follows: ethanol concentration, 60%; extraction pressure, 400 MPa; extraction time, 2 min; extraction temperature, 30 °C; and the solid/liquid ratio, 1 : 50. Under these conditions, the yields of chlorogenic acid and cynaroside were raised to 4.863% and 0.080%, respectively. Compared with other extraction methods, such as heat reflux extraction (HRE), ultrasonic extraction (UE), and Sohxlet extraction (SE), the UPE method showed several advantages, including higher extraction yield, shorter extraction time, lower energy consumption, and higher purity of the extracts. This study could help better utilize L. japonica flower buds as a readily accessible source of natural antioxidants in food and pharmaceutical industries.

Variations in intraplatelet phospho-VASP expression due to pre-analytical sample preparations, illustration of a quality control issue in platelet pharmacology

Intraplatelet vasodilator-stimulated phosphoprotein [VASP] analysis is a commonly used laboratory approach for monitoring of the anti-platelet therapy with adenosine diphosphate [ADP] receptor blocking agents; however, it's testing in clinical laboratory needs a high level of experience and proficiency. The ability to recognize how the pre-analytical variations can change the results would be helpful for the interpretation of data from intraplatelet VASP analysis. The aim of this study was to describe the possible differences of intraplatelet phospho- VASP expression between washed and platelet rich plasma [PRP] samples, both at baseline levels and following experimentally induction of VASP phosphorylation. PRP and washed platelet samples were treated with different inducers of VASP phosphorylation, including forskolin [10 micro M], prostaglandin E1 [PGE1] [50 nM] and sodium nitro-prusside [SNP] [100 micro M]. Untreated PRP and washed platelet samples were also included in study as baseline controls. After labeling of platelets with either anti P-Serine157-VASP or anti P-Serine239-VASP, the samples were subjected to flow cytometric analysis to monitor the levels of intraplatelet phospho-VASP expression. Washed platelet samples tend to show increased expression of intraplatelet P-Serine[157]- VASP at baseline state and also more expression of P-Serine[157]-VASP and P-Serine[239]-VASP in response to forskolin and SNP, compared with PRP samples. Though, reduced levels of PGE1- induced VASP phosphorylation at both residues were detected for washed platelets. In this study we have provided some background information required for performing of intraplatelet VASP analysis on differently handled platelet samples and interpretation of the obtained results

Pruebas bioquímicas tradicionales y de alta resolución para identificación manual de enterobacterias / Traditional and high-resolution biochemical tests for manual identification of enterobacteria / Provas bioquimicas tradicionais e de alta resolução para identificação manual de enterobacterias

Se comparó una serie tradicional de pruebas bioquímicas con una de alta resolución para identificar 500 cepas de enterobacterias utilizando un método probabilístico para interpretar los resultados. La serie tradicional estuvo formada por 10 pruebas (producción de ornitina descarboxilasa, lisina descarboxilasa, lisina desaminasa, ácido sulfhídrico, indol y gas, hidrólisis de urea, utilización de citrato y de malonato, y movilidad). La serie de alta resolución, también con 10 pruebas, se integró con las primeras 4 mencionadas en la serie tradicional y 6 de fermentación de hidratos de carbono (adonitol, L-arabinosa, celobiosa, L-ramnosa, rafinosa y sorbitol). Con la serie de alta resolución se asignaron identidades únicas a 445 cepas (351 con probabilidad de 1,0 y 94 con probabilidades entre 0,010 y 0,999), y de las restantes 55 cepas, a 53 y 2 se asignaron dos y tres identidades probables respectivamente. Con la serie tradicional se asignaron identidades únicas a 306 cepas (110 con probabilidad de 1,0 y 196 con probabilidades entre 0,001 y 0,999) y a 179 y 5 se asignaron dos y tres identidades probables respectivamente. Diez cepas no se pudieron identificar. Todos los indicadores analizados revelaron la superioridad de la serie de alta resolución. El método probabilístico permitió la comparación objetiva de ambas series.

A traditional series of biochemical tests-was compared to a high-resolution one in order to identify 500 strains of enterobacteria, using a probabilistic method for the interpretation of experimental results. The traditional series was formed by 10 tests (ornithine decarboxylase, lysine decarboxylase, lysine deaminase, sulfhydric acid, indol and gas production, urea hydrolysis, citrate and malonate utilization, and motility). The high-resolution one was also formed by 10 tests, including the first 4 tests mentioned above and 6 carbohydrate fermentation tests (adonitol, L-arabinose, cellobiose, L-rhamnose, raffinose, and sorbitol). With the high-resolution series, single identities were assigned to 445 strains (351 with a probability of 1.0 and 94 with probabilities in the range 0.010-0.999), and for the remaining strains two and three probable identities were assigned to 53 and 2 strains, respectively. With the traditional series, single identities were assigned to 306 strains (110 with a probability of 1.0 and 196 with probabilities in the 0.001-0.999 range), two and three probable identities were assigned to 179 and 5 strains respectively; 10 strains turned out to be non-identifiable. Every parameter of comparison used revealed the superiority of the high-resolution series. The probabilistic method for interpretation of experimental results allowed an objective comparison of both series.

Foi comparada uma série tradicional de testes bioquímicos com uma outra de alta resolução para identificar 500 cepas de enterobactérias usando uma abordagem probabilística para a interpretação dos resultados. A série tradicional consistiu em 10 testes (produção de ornitina descarboxilase, lisina descarboxilase, lisina desaminase, ácido sulfídrico, indol e gás, hidrólise da ureia, utilização de citrato e de malonato, e mobilidade). A série de alta resolução, também com 10 ensaios, integrou-se com as primeiras 4 mencionadas na série tradicional e 6 de fermentação de hidratos de carbono (adonitol, L-arabinose, celobiose, L-ramnose, rafinose e sorbitol). Com a série de alta resolução foram atribuídas identidades únicas a 445 cepas (351, com probabilidade 1,0, e 94, com probabilidade entre 0,010 e 0,999), e das restantes 55 cepas, a 53 e 2 foram atribuídas duas e três identidades possíveis, respectivamente. Com a série tradicional foram atribuídas identidades únicas a 306 cepas (110 com probabilidade de 1,0 e 196 com probabilidades entre 0,001 e 0,999) e a 179 e 5 foram atribuídas duas e três identidades prováveis, respectivamente. Dez cepas não puderam ser identificadas. Todos os indicadores analisados demonstraram a superioridade da série de alta resolução. O método probabilístico permitiu a comparação objetiva de ambas as séries.

Development and validation of analytical method for determining the artificial dyes in breakfast cereals by means of high performance liquid chromatography

Validation of analytical methodology is an important tool to ensure the applicability and scope ofa technique for laboratory routine, establishing the limits of the quality parameters of instrumental measurements and the statistical reliability by estimating the procedure performance. There are several normative documents for establishing the figures of merit, namely, limit of detection (LOD), limit of quantification (LOQ), linearity, selectivity, repetitivity, intermediate precision and recovery. This workaimed at developing and validating an analytical methodology for quantifying the artificial dyes in cereal by means of high performance liquid chromatography (HPLC). All of the validation process of this study was performed according to the recommendations by Thompson et al, and following the guidance statedin the document INMETRO DOQ-CGCRE-08 and the harmonized Guidelines IUPAC. After completing all of the validation steps, the methodology showed to be precise, exact, linear over a wide concentration range and its analysis has not being influenced by the food matrix for HPLC determination. More over, the methodology has shown as an important contribution since no official methodologies have been available for determining artificial dyes in breakfast cereals...

Preparación intestinal colónica con polietilenglicol y manitol: efectividad según la escala de Boston / Colonic bowel preparation: effectiveness of polyethyleneglycol and manitol

Comparar efectividad del polietilenglicol y manitol en la preparación intestinal mediante escala de Boston, pacientes de la consulta externa de gastroenterología, tercer trimestre, 2012. Estudio prospectivo, transversal, experimental. Muestra de 100 pacientes aleatorizados en dos grupos: polietilenglicol y manitol, 50 en cada uno. A todos se les instauró dieta líquida el día previo al estudio e indicación para la ingesta de la solución a evaluar. Se realizó colonoscopia con evaluación endoscópica según escala de Boston. La tolerancia a la preparación fue considerada fácil por 88% en el grupo polietilenglicol vs 100% del grupo manitol (p=0,041). El 98% del grupo manitol consideró que este medicamento tenía sabor agradable en comparación con polietilenglicol (78%) (p=0,002). El efecto adverso más frecuente en ambos grupos fue la náusea. El polietilenglicol alcanzó exploraciones completas con restos en un 82% colon derecho, 56% colon transverso y 72% colon izquierdo, mientras que con manitol prevaleció la exploración completa sin restos en 66%, 90% y 68% respectivamente (p<0,05). La puntuación global de la escala de Boston con polietilenglicol y manitol fue 6 vs 8 (p<0,05). Manitol resultó ser más efectivo que polietilenglicol para la preparación del colon en su totalidad y por segmentos

To compare the effectiveness of polyethyleneglycol and mannitol bowel preparation by Boston scale, in patients from the outpatient gastroenterology in the third quarter of 2012. Prospective, cross, experimental with a sample of 100 patients randomized to group polyethyleneglycol and mannitol group, 50 in each. All were introduced liquid diet the day before the test with the appropriate indication for the intake of the solution to evaluate and colonoscopy was performed endoscopic evaluation scale as Boston. Tolerance was considered easy preparation by 88% in polyethyleneglycol group vs 100% mannitol group (p=0.041). 98% mannitol group had considered that this medicine palatable compared with polyethyleneglycol (78%) (p=0.002). The most common adverse event in both groups was nausea. Polyethyleneglycol reached full scans with remains at 82% right colon, transverse colon 56% and 72% left colon, whereas mannitol prevailed without full exploration remains at 66%, 90% and 68% respectively (p<0,05). The overall rating scale was polyethyleneglycol Boston 6 vs 8 in the mannitol group (p<0,05). Mannitol was more effective for the preparation of polyethyleneglycol entire colon and segments

Sample pretreatment methods of pesticide residues in Panax notoginseng of Chinese traditional medicine / 药学学报

Two sample pretreatment methods of pesticide residues in Panax notoginseng of Chinese traditional medicine were developed. For Method I, the residues were extracted from homogenized tissue with n-hexane-dichloromethane (6:4) by means of ultrasonication, the crude extract was purified by an Envi-carb/NH2 solid-phase extraction (SPE) column. For Method II, matrix solid-phase dispersion (MSPD) technique was used for extracting and cleaning up. The eluates were concentrated by rotary evaporation, and then were redissolved in dichloromethane prior to GC-MS determination. The determination was performed in selected ion monitoring (SIM) mode with the external calibration for quantitative analysis. Under the optimal conditions, the results indicated that the methods are easier and faster, the recoveries of method I for the spiked standards at concentration of 0.01, 0.5, and 2.0 mg x kg(-1) were 81.90%-102.10% with the relative standard deviations (RSDs) of 3.60%-7.10%. The recoveries of method II were 96.26%-104.20% with the RSDs of 3.52%-7.94%. The detection limits (S/N) for residues of pesticides were in the range of 0.48-1.34 ng x g(-1). The results indicated that these multiresidue analysis methods can meet the requirements for determination of residue pesticides and can be appropriate for trace analysis of residue pesticides in Panax notoginseng.

Chromatographic methods for biogenic amines determination in foods of animal origin / Métodos cromatográficos para determinar aminas biogênicasem alimentos de origem animal

Biogenic amines (BAs) are formed as a result of specific free amino acid decarboxylation. Analysis of these metabolitesmay be of great importance to determine food quality and for monitoring the levels of biogenic amines such as histamineand tyramine related to intoxication episodes in humans. Chromatography is a chemistry separation technique usedto characterize biogenic amines in foods. Variations of this technique (liquid, thin layer and gas chromatography)have been widely applied; however, the food matrix complex requires that changes in the methodology of extraction,derivatization and detection must be performed according to each group of foods. High-performance liquidchromatography is the most widely used chromatographic method applied for biogenic amines in foods. However, dueto the current importance of biogenic amines in quality control and consumer safety, researchers try to develop newmethods for a fast, reliable analysis of foods in the market. This review presents some chromatographic techniquesapplied to monitoring BAs in different foods of animal origin...

Aminas biogênicas são formadas como resultado da descarboxilação de aminoácidos livres específicos. A análise dessesmetabólitos é de grande importância na determinação da qualidade e monitoramento de biogênicas como histamina etiramina relacionadas com episódios de intoxicação em humanos. A cromatografia é uma técnica de separação químicausada para caracterizar aminas biogênicas. Variações da técnica (cromatografia líquida, em camada delgada e gasosa)têm sido amplamente usadas, porém a complexidade da matriz alimentar faz com que sejam realizadas mudanças nosprocessos de extração, derivatização e detecção em concordância com cada grupo de alimento. A cromatografia líquidade alta eficiência (CLAE) é o método mais utilizado na determinação de aminas biogênicas em alimentos. Contudo,devido à importância das aminas biogênicas no controle da qualidade e a segurança do consumidor, os pesquisadorestentam desenvolver novos métodos com o intuito de uma análise mais rápida e precisa para o controle de alimentos nomercado. O objetivo da revisão é apresentar algumas técnicas cromatográficas aplicadas no monitoramento de aminasbiogênicas em produtos de origem animal...

Rapid titrimetric and spectrophotometric determination of ofloxacin in pharmaceuticals using N-bromosuccinimide / Determinação titulométrico e espectrofotométrico rápido de ofloxacina em produtos farmacêuticos usando N -bromosuccinimida

One titrimetric and two spectrophotometric methods have been described for the determination of ofloxacin (OFX) in bulk drug and in tablets, employing N-Bromosuccinimide as an analytical reagent. The proposed methods involve the addition of a known excess of NBS to OFX in acid medium, followed by determination of unreacted NBS. In titrimetry, the unreacted NBS is determined iodometrically, and in spectrophotometry, unreacted NBS is determined by reacting with a fixed amount of either indigo carmine (Method A) or metanil yellow (Method B). In all the methods, the amount of NBS reacted corresponds to the amount of OFX. Titrimetry allows the determination of 1-8 mg of OFX and the calculations are based on a 1:5 (OFX:NBS) reaction stoichiometry. In spectrophotometry, Beer's law is obeyed in the concentration ranges 0.5-5.0 µg/mL for method A and 0.3-3.0 µg/mL for method B. The molar absorptivities are calculated to be 5.53x10(4) and 9.24x10(4) L/mol/cm for method A and method B, respectively. The methods developed were applied to the assay of OFX in tablets, and results compared statistically with those of a reference method. The accuracy and reliability of the methods were further ascertained by performing recovery tests via the standard-addition method.

Descrevem-se métodos, um titulométrico e dois espectrofotométricos, para a determinação de ofloxacino (OFX) na matéria-prima e em comprimidos, empregando a N-bromossuccinimida (NBS) como reagente analítico. Os métodos propostos envolvem a adição de excesso conhecido de NBS ao OFX, em meio ácido, seguida de determinação do NBS que não reagiu. Na titulometria, o NBS que não reagiu é determinado iodometricamente e na espectrofotometria, o NBS que não reagiu é determinado pela reação com quantidade fixa de índigo carmim (Método A) ou amarelo de metanila (Método B). Em todos os métodos, a quantidade de NBS que reagiu corresponde à quantidade de OFX. A titulometria permite a determinação de 1-8 mg de OFX e os cálculos se baseiam na estequiometria de reação de 1:5 (OFX:NBS). Na espectrofotometria, a Lei de Beer é obedecida nas faixas de concentração de 0,5-5,0 µg/mL, para o método A, e de 0,3-3,0 µg/mL, para o método B, respectivamente. Os métodos desenvolvidos foram aplicados para o teste de OFX em comprimidos e os resultados foram comparados estatisticamente com aqueles do método de referência. A precisão e a confiabilidade dos métodos foram, posteriormente, verificadas por meio dos testes de recuperação via método de adição de padrão.

Factores ambientales y genéticos asociados al cáncer de mama en féminas del área de salud 28 de septiembre / Environmental and genetic factors associated with breast cancer in women from 28 de September health area

El cáncer de mama ocupa, tanto por el número de pacientes diagnosticadas como fallecidas por esta causa, uno de los primeros lugares en el mundo y también en Cuba; razones estas que justificaron la realización de un estudio analítico observacional de casos y controles en el área de salud 28 de Septiembre de Santiago de Cuba durante el cuatrimestre enero-abril de 2009, que incluyó a 40 mujeres con cáncer mamario (consideradas como casos), registradas en el Servicio de Patología de Mama y seleccionadas mediante un muestreo aleatorio simple, así como a 80 controles sin este diagnóstico, para determinar los posibles factores ambientales y genéticos que pudieron haber influido en la aparición de esta neoplasia. Se aplicó la prueba de Ji al cuadrado, con un nivel de significación de 0,05 y se calculó la oportunidad relativa (odds ratio) para evaluar la magnitud de asociación entre variables y por intervalo de confianza. Se halló asociación de antecedentes patológicos familiares de ese tipo de cáncer en los casos, sobre todo en parientes de primer grado y más significativo en el grupo con la enfermedad, en el que también se observó más comúnmente agregación familiar de la afección. Se concluyó que esa formación neoplásica fue más frecuente en mujeres de 51 a 65 años, con menopausia tardía y hábitos tóxicos.

Breast cancer, for the number of diagnosed patients and those dead by this cause, is in one of the first positions in the world and also in Cuba; reasons why an analytic observational case-control study in 28 de Septiembre health area of Santiago de Cuba was carried out from January to April 2009, which included 40 women with breast cancer (considered as cases), who were recorded in the Service of Breast Pathology and selected by a simple random sampling, as well as 80 controls without this diagnosis, to determine possible environmental and genetic factors influencing the occurrence of this malignancy. The chi-square test was performed with a significance of 0,05, and odds ratio was calculated to evaluate the extent of association between variates and by confidence interval. An association of family medical history of this type of cancer was found in the cases, particularly in first-degree relatives and more significantly in the group with this disease, in which disease family aggregation was also observed more commonly. It is concluded that this malignancy was more frequent en women between 51 and 65 years with late menopause and toxic habits.

HPLC method development for the simultaneous analysis of amlodipine and valsartan in combined dosage forms and in vitro dissolution studies

A simple, rapid and reproducible HPLC method was developed for the simultaneous determination of amlodipine and valsartan in their combined dosage forms, and for drug dissolution studies. A C18 column (ODS 2, 10 μm, 200 x 4.6 mm) and a mobile phase of phosphate buffer (pH 3.6 , 0.01 mol L-1):acetonitrile: methanol (46:44:10 v/v/v) mixture were used for separation and quantification. Analyses were run at a flow-rate of 1 mL min-1 and at ambient temperature. The injection volume was 20 μL and the ultraviolet detector was set at 240 nm. Under these conditions, amlodipine and valsartan were eluted at 7.1 min and 3.4 min, respectively. Total run time was shorter than 9 min. The developed method was validated according to the literature and found to be linear within the range 0.1 - 50 μg mL-1 for amlodipine, and 0.05 - 50 μg mL-1 for valsartan. The developed method was applied successfully for quality control assay of amlodipine and valsartan in their combination drug product and in vitro dissolution studies.

Desenvolveu-se método de HPLC rápido e reprodutível para a determinação simultânea de anlodipino e valsartana em suas formas de associação e para os estudos de dissolução dos fármacos. Utilizaram-se coluna C18 (ODS 2, 10 μm, 200 x 4,6 mm) e fase móvel tampão fosfato (pH 3,6, 0,01 mol L-1):acetonitrila: metanol para a separação e a quantificação. As análises foram efetuadas com velocidade de fluxo de 1 mL min-1 e à temparatura ambiente O volume de injeção foi de 20 μL e utilizou-se detector de ultravioleta a 240 nm. Sob essas condições, anlodipino e valsartana foram eluídas a 7,1 min e 3,4 min, respectivamente. O tempo total de corrida foi menor que 9 min. O método desenvolvido foi validado de acordo com a literatura e se mostrou linear na faixa de 0,1-50 μg mL-1 para anlodipino e de 0,05-50 μg mL-1 para valsartana. O método desenvolvido foi aplicado com sucesso para ensaios de controle de qualidade de associações de anlodipino e valsartana e nos estudos de dissolução in vitro.

Estimação por aproximação química dos teores de proteína indegradável insolúvel em detergente neutro em forragens tropicais / Estimation of undegradable neutral detergent insoluble protein in tropical forages by chemical approaching

A mathematical model was developed to predict the content of undegradable neutral detergent insoluble protein (UNDIP) from chemical characteristics of tropical forages. This study was based on a biological limitation of a previous model, which restricts the UNDIP estimates to values equals or higher than 1.34 percent of dry matter. The databank was formed by 540 samples of tropical forages used in cattle feeding (fresh forage and hay). The ratio of UNDIP on neutral detergent insoluble protein (NDIP) was chosen as response variable and the acid detergent insoluble protein (ADIP) as independent variable. The mathematical model was found to be exponential, assuming the formula: UNDIP = NDIP x e-(0,818+0.16764DIP) , in which all values are expressed on dry matter basis. It was observed that biological limitation of the previous model was eliminated, even though a low statistical improvement was obtained. The prediction of a biological parameter (UNDIP) from a chemical characteristic (ADIP) still have some restrictions and the estimates should be applied with caution in some situations. The main application of the model described above is estimate UNDIP contents when biological methods are not available.