Development of a validated stability-indicating HPLC- DAD method for dasabuvir and the characterization of its degradation products using LC-QToF-MS/MS

Abstract A stability-indicating HPLC-DAD method was developed and validated for the simultaneous determination of dasabuvir and its degradation products in the pharmaceutical formulation. The proposed method utilized a Symmetry® C18 (4.6 x 75 mm, 3.5 µm) column, and the mobile phase consisted of an isocratic elution of formic acid (0.1%) and acetonitrile (55:45, v/v), at a flow of 1 mL min-1; analytes were detected at 244 nm. Dasabuvir was submitted to different stress degradation conditions, such as acidic, alkaline, neutral, thermal, oxidative and photolytic, and the structural elucidation of degradation products was performed using LC-QToF-MS/MS. The HPLC-DAD stability-indicating method was validated for selectivity, linearity, limit of detection and quantification, accuracy, precision and robustness, according to ICH guidelines. Dasabuvir produced two degradation products (DP1 and DP2) from the alkaline stress conditions, which were characterized in negative ion mode. Dasabuvir was linear in the range 9.78 to 136.92 µg mL-1, and DP and DP were linear in the range 2.9 to 20.2 µg mL-1 and 1.3 to 14.9 µg mL-1, respectively. The 1 2 recovery ranged between 99.16 and 100.86%, while precision ranged from 1.02 to 2.89%. As the method can effectively separate the dasabuvir from its degradation products and quantitate them, it may be employed as a stability-indicating method for the pharmaceutical formulation.

A multivariate analysis of risk factors associated with death by Covid-19 in the USA, Italy, Spain, and Germany.

Aim: Our aim was to investigate the risk factors associated with death from COVID-19 in four countries: The USA, Italy, Spain, and Germany. Subject and methods: We used data from the Institute for Health Metrics and Evaluation with projection information from January-August 2020. A multivariate analysis of logistic regression was performed. The following factors were analyzed (per day): number of beds needed for the hospital services, number of intensive care units (ICU) beds required, number of ventilation devices, number of both hospital and ICU admissions due to COVID-19. Nagelkerke's R2 coefficient of determination was used to evaluate the model's predictive ability. The quality of the model's fit was assessed by the Hosmer-Lemeshow and the chi-square tests. Results: Among the evaluated countries, Italy presented greater need for ICU beds/day (≤ 98; OR = 2315.122; CI 95% [334.767-16,503.502]; p < 0.001) and daily ventilation devices (≤ 118; OR = 1784.168; CI 95% [250.217-12,721.995]; p < 0.001). It is expected that both Italy and Spain have a higher ICU admission rate due to COVID-19 (n = 14/day). Spain will need more beds/day (≤ 357; OR = 146.838; CI 95% [113.242-190.402]; p  < 0.001) and probably will have a higher number of daily hospital admissions (n = 48/day). All the above-mentioned factors have an important impact on patients' mortality due to COVID-19 in all four countries. Conclusions: Further investments in hospitals' infrastructure, as well as the development of innovative devices for patient's ventilation, are paramount to fight the pandemic in the USA, Italy, Spain, and Germany.

Risk factors associated with delay in diagnosis and mortality in patients with COVID-19 in the city of Rio de Janeiro, Brazil.

We investigated the predictors of delay in the diagnosis and mortality of patients with COVID-19 in Rio de Janeiro, Brazil. A cohort of 3,656 patients were evaluated (Feb-Apr 2020) and patients' sociodemographic characteristics, and social development index (SDI) were used as determinant factors of diagnosis delays and mortality. Kaplan-Meier survival analyses, time-dependent Cox regression models, and multivariate logistic regression analyses were conducted. The median time from symptoms onset to diagnosis was eight days (interquartile range [IQR] 7.23-8.99 days). Half of the patients recovered during the evaluated period, and 8.3% died. Mortality rates were higher in men. Delays in diagnosis were associated with male gender (p = 0.015) and patients living in low SDI areas (p < 0.001). The age groups statistically associated with death were: 70-79 years, 80-89 years, and 90-99 years. Delays to diagnosis greater than eight days were also risk factors for death. Delays in diagnosis and risk factors for death from COVID-19 were associated with male gender, age under 60 years, and patients living in regions with lower SDI. Delays superior to eight days to diagnosis increased mortality rates.

Risk factors associated with delay in diagnosis and mortality in patients with COVID-19 in the city of Rio de Janeiro, Brazil / Fatores de risco associados ao atraso no diagnóstico e mortalidade em pacientes com COVID-19 na cidade do Rio de Janeiro, Brasil

Abstract We investigated the predictors of delay in the diagnosis and mortality of patients with COVID-19 in Rio de Janeiro, Brazil. A cohort of 3,656 patients were evaluated (Feb-Apr 2020) and patients' sociodemographic characteristics, and social development index (SDI) were used as determinant factors of diagnosis delays and mortality. Kaplan-Meier survival analyses, time-dependent Cox regression models, and multivariate logistic regression analyses were conducted. The median time from symptoms onset to diagnosis was eight days (interquartile range [IQR] 7.23-8.99 days). Half of the patients recovered during the evaluated period, and 8.3% died. Mortality rates were higher in men. Delays in diagnosis were associated with male gender (p = 0.015) and patients living in low SDI areas (p < 0.001). The age groups statistically associated with death were: 70-79 years, 80-89 years, and 90-99 years. Delays to diagnosis greater than eight days were also risk factors for death. Delays in diagnosis and risk factors for death from COVID-19 were associated with male gender, age under 60 years, and patients living in regions with lower SDI. Delays superior to eight days to diagnosis increased mortality rates.

Resumo Investigamos os preditores de atraso no diagnóstico e mortalidade de pacientes com COVID-19 no Rio de Janeiro, Brasil. Uma coorte de 3.656 pacientes foi avaliada (fevereiro-abril de 2020) e as características sociodemográficas dos pacientes, o bairro e o índice de desenvolvimento social (IDS) foram usados como fatores determinantes dos atrasos no diagnóstico e da mortalidade. Foram realizadas análises de sobrevivência de Kaplan-Meier, modelos de regressão Cox dependentes do tempo e análises de regressão logística multivariada. O tempo mediano desde o início dos sintomas até o diagnóstico foi de oito dias (intervalo interquartil [IQR] 7,23-8,99 dias). Metade dos pacientes se recuperou no período avaliado e 8,3% faleceram. As taxas de mortalidade foram maiores nos homens. Atrasos no diagnóstico foram associados ao sexo masculino (p = 0,015) e pacientes que moravam em áreas com baixo IDS (p < 0,001). As faixas etárias estatisticamente associadas à morte foram: 70-79 anos, 80-89 anos e 90-99 anos. Atrasos no diagnóstico superiores a oito dias também foram fatores de risco para óbito. Atrasos no diagnóstico e fatores de risco para morte por COVID-19 foram associados ao sexo masculino, idade abaixo de 60 anos e pacientes que vivem em regiões com menor IDS. Atrasos superiores a oito dias no diagnóstico aumentam as taxas de mortalidade.

Morpho-anatomy of the inflorescence of Musa × paradisiaca

ABSTRACT Bananas and plantains are herbaceous monocotyledonous plants belonging to the genus Musa, Musaceae, which has a widespread distribution around the world. Various parts of banana plant are commonly used in traditional medicines. Several species of Musa are reported to possess anti-inflammatory, anti-hyperglycemic and antidiabetic properties. This work is aimed at studying the morphological and anatomical characteristics of the inflorescences of Musa × paradisiaca L., that could contribute to the characterization of these species cultivated in Brazil. Plant materials were collected and prepared in accordance with standard optical microscopy techniques. Morphological characterizations were conducted using morphological descriptors for inflorescences, including some descriptors from International Plant Genetic Resources Institute for Musa spp. Microscope slides were prepared using glycol-methacrylate and were stained in toluidine blue. Main features observed for M. × paradisiaca inflorescence were amphistomatic bracts with tetracytic stomata, fiber caps next to the phloem, adaxial and abaxial uniseriate epidermis, and papillose on the abaxial face. Outer tepals have multilayer epidermis and vascular bundles aligned next to the abaxial face. Free tepal has unilayeredepidermis. Anthers are tetrasporangiate and the locules are separated by the septum. Ovary is inferior and trilocular with external unilayered and internal epidermis. The main morpho-anatomical characteristics of inflorescence of Musa × paradisiaca are highlighted in this study, contributing to provide more information about the characterization of this species cultivated in Brazil.

A new HILIC-MS/MS method for the simultaneous analysis of carbidopa, levodopa, and its metabolites in human plasma.

Monitoring of the plasmatic levels of levodopa (LEV) and carbidopa (CAR) is necessary to adjust the dose of these drugs according to the individual needs of Parkinson's disease patients. To support drug therapeutic monitoring, a method using HILIC mode and LC-MS/MS was developed for the simultaneous determination of carbidopa, levodopa, and its metabolites (3-o-methyldopa (3-OMD) and dopamine (DOPA)) in human plasma. A triple quadrupole mass spectrometry was operated under the multiple reaction-monitoring mode (MRM) using the electrospray ionization technique. After straightforward sample preparation via protein precipitation, an Atlantis HILIC (150 × 2.1 mm, 3 µm, Waters, USA) column were used for separation under the isocratic condition of acetonitrile/water (79:21, v/v) containing 0.05% formic acid and 3 mmol/L ammonium formate and the total run time was 7 min. Deuterated LEV was used as internal standard for quantification. The developed method was validated in human plasma with a lower limit of quantitation of 75 ng/mL for LEV, 65 ng/mL for CAR and 3-OMD, and 20 ng/mL for DOPA. The calibration curve was linear within the concentration range of 75-800 ng/mL for LEV, 65-800 ng/mL for CAR and 3-OMD, and 20-400 ng/mL for DOPA (r>0.99). The assay was accurate and precise, with inter-assay and intra-assay accuracies within ±13.44% of nominal and inter-assay and intra-assay precision≤13.99%. All results were within the acceptance criteria of the US FDA and ANVISA guidelines for method validation. LEV, CAR, 3-OMD and DOPA were stable in the battery of stability studies, long-term, bench-top, autosampler, and freeze/thaw cycles. Samples from patients undergoing treatment were analyzed, and the results indicated that this new method is suitable for therapeutic drug monitoring in Parkinson's disease patients.