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.

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

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.