Identification of Asiaticoside from Centella erecta (Apiaceae) as Potential Apyrase Inhibitor by UF-UHPLC-MS and Its In Vivo Antischistosomal Activity.

Schistosomiasis, caused by parasites of the genus Schistosoma, is a neglected disease with high global prevalence, affecting more than 240 million people in several countries. Praziquantel (PZQ) is the only drug currently available for the treatment. S. mansoni NTPDases (known as SmNTPDases, ATP diphosphohydrolases or ecto-apyrases) are potential drug targets for the discovery of new antischistosomal drugs. In this study, we screen NTPDases inhibitors from Centella erecta (Apiaceae) using an ultrafiltration combined UHPLC-QTOF-MS method and potato apyrase, identifying asiaticoside as one of the apyrase-binding compounds. After isolation of asiaticoside from C. erecta extract, we assessed its in vivo antischistosomal activities against Schistosoma mansoni worms and its in vitro enzymatic apyrase inhibition. Also, molecular docking analysis of asiaticoside against potato apyrase, S. mansoni NTPDases 1 and 2 were performed. Asiaticoside showed a significant in vitro apyrase inhibition and molecular docking studies corroborate with its possible actions in potato apyrase and S. mansoni NTPDases. In mice harboring a patent S. mansoni infection, a single oral dose of asiaticoside (400 mg/kg. p.o.) showed significantly in vivo antischistosomal efficacy, markedly decreasing the total worm load and egg burden, giving support for further exploration of apyrase inhibitors as antischistosomal agents.

Simultaneous Monitoring of Azathioprine Metabolites in Erythrocytes of Crohn's Disease Patients by HPLC-UV.

The conversion of azathioprine (AZA) to active 6-thioguanine nucleotides (6-TGN) is essential for its clinical efficacy; however, another metabolite formed, 6-methylmercaptopurine (6-MMP), is related to hepatotoxicity. Blood samples were collected from 37 patients under AZA's treatment, and a new HPLC-UV method was validated and applied for simultaneous quantification of 6-TGN and 6-MMP in erythrocytes of Crohn's disease (CD) patients. The concentration of 6-TGN and 6-MMP found ranged from 4.5 to 2,456 ρmol/8 × 108 red blood cells (RBCs) for 6-TGN and from 170 to 53,951 ρmol/8 × 108 RBCs for 6-MMP. Reduced levels of 6-MMP in patients into combo therapy with AZA and allopurinol (2,031 ρmol/8 × 108 RBCs) have been observed when compared to patients undergoing monotherapy with AZA (9,098 ρmol/8 × 108 RBCs). Additionally, there was a negative correlation (r = -83.7%, P < 0.05) between lymphocyte count and 6-TGN levels. The method developed is reliable, accurate and reproducible and can be used as an important tool in the monitoring routine of patients with CD under AZA treatment, allowing the individualization of the dose, monitoring adherence to the treatment and the evaluation of the clinical outcome of these patients.

Nebulization of Vancomycin Provides Higher Lung Tissue Concentrations than Intravenous Administration in Ventilated Female Piglets with Healthy Lungs.

BACKGROUND: Intravenous vancomycin is used to treat ventilator-associated pneumonia caused by methicillin-resistant Staphylococcus aureus, but achieves high rates of failure. Vancomycin nebulization may be efficient to provide high vancomycin lung tissue concentrations. The aim of this study was to compare lung tissue and serum concentrations of vancomycin administered intravenously and by aerosol in mechanically ventilated and anesthetized healthy piglets. METHODS: Twelve female piglets received a single intravenous dose of vancomycin (15 mg/kg) and were killed 1 (n = 6) or 12 h (n = 6) after the end of administration. Twelve piglets received a single nebulized dose of vancomycin (37.5 mg/kg) and were killed 1 (n = 6) or 12 h (n = 6) after the end of the aerosol administration. In each group, vancomycin lung tissue concentrations were assessed on postmortem lung specimens using high-performance liquid chromatography. Blood samples were collected for serum vancomycin concentration measurement 30 min and 1, 2, 4, 6, 8, and 12 h after the end of vancomycin administration. Pharmacokinetics was analyzed by nonlinear mixed effect modeling. RESULTS: One hour after vancomycin administration, lung tissue concentrations in the aerosol group were 13 times the concentrations in the intravenous group (median and interquartile range: 161 [71, 301] µg/g versus 12 [4, 42] µg/g; P < 0.0001). Twelve hours after vancomycin administration, lung tissue concentrations in the aerosol group were 63 (23, 119) µg/g and 0 (0, 19) µg/g in the intravenous group (P < 0.0001). A two-compartment weight-scaled allometric model with first-order absorption and elimination best fit serum pharmacokinetics after both routes of administration. Area under the time-concentration curve from 0 to 12 h was lower in the aerosol group in comparison to the intravenous group (56 [8, 70] mg · h · l vs. 121 [103, 149] mg · h · l, P = 0.002). Using a population model, vancomycin bioavailability was 13% (95% CI, 6 to 69; coefficient of variation = 85%) and absorption rate was slow (absorption half life = 0.3 h). CONCLUSIONS: Administration of vancomycin by nebulization resulted in higher lung tissue concentrations than the intravenous route.

Comparação entre métodos de diagnósticos para a quantificação da lipoproteína de baixa densidade em pacientes atendidos em um Hospital Universitário / Comparison between diagnostic methods for the quantification of low-density lipoprotein in patients attended at a University Hospital

Introdução: Estudos relatam uma forte ligação entre as dislipidemias e as ateroscleroses. Por esta razão, exames como o perfil lipídico são realizados rotineiramente com o intuito de prevenção e monitoramento dessas doenças. A lipoproteína de baixa densidade possui grande destaque por apresentar maior relação com fatores de risco para o desenvolvimento de doenças ateroscleróticas. Métodos diretos de obtenção dos valores dessa lipoproteína são confrontados com resultados obtidos usualmente na rotina, através de equações que fornecem valores estimados. Objetivo: Comparar os métodos de diagnósticos utilizados para a obtenção da lipoproteína de baixa densidade através das Fórmulas de Friedewald e Martin com os resultados obtidos por metodologia automatizada, em pacientes atendidos em um Hospital Universitário de Juiz de Fora - MG. Material e Métodos: Foram coletadas amostras de sangue venoso para a obtenção do soro de 208 pacientes. Quantificaram-se os níveis de colesterol total, triglicerídeos e da lipoproteína de alta densidade para a obtenção da lipoproteína de baixa densidade através das equações de Friedewald e Martin. Resultados: Há uma correlação positiva entre os resultados de lipoproteína de baixa densidade calculados pelos métodos de Martin e direto (ρ=0,916), e uma correlação positiva entre os resultados pelos métodos de Friedewald e direto (ρ=0,915). Discussão: Foi observada uma correlação positiva entre os valores de colesterol e de lipoproteína de baixa densidade pelas três metodologias. O método de Bland-Altman foi utilizado para comparação dos resultados obtidos pelas equações e pela metodologia direta. Conclusão: Ainda que as equações de Friedewald e Martin tenham apresentado boa correlação com a lipoproteína de baixa densidade medida por metodologia direta, estudos que relacionam doenças arteriais ateroscleróticas à lipoproteína de baixa densidade devem considerar a quantificação direta desta a fim de abranger os indivíduos com suas diversas especificidades.

Introduction: Studies have reported a strong link between dyslipidemia and atherosclerosis. For this reason, exams such as the lipid profile are routinely performed for the prevention and monitoring of these diseases. Among the lipid indices, low density lipoproteins should be highlighted because due to their greater relation with risk factors for the development of atherosclerotic diseases. Therefore, direct methods of obtaining low density lipoproteins values, considered more accurate, are confronted with results usually obtained in the routine, through equations that provide estimated values. Objective:We compared the diagnostic methods used to obtain low density lipoproteins through the Friedewald and Martin formulas with the results obtained by automated methodology in patients attended at a University Hospital of Juiz de Fora ­ MG. MaterialandMethods: A total of 208 patients were recruited and venous blood samples were collected to obtain serum. The levels of total cholesterol, triglycerides and high density lipoprotein were quantified to obtain low density lipoproteins through the Friedewald and Martin equations. Results:A positive correlation between low density lipoproteins results has calculated by Martin and direct methods (ρ = 0.916), and positive correlation between Friedewald results and direct (ρ = 0.915). Discussion: We observed a positive correlation between the values of cholesterol and low density lipoproteins by the three methodologies. The Bland-Altman method has been used to compare the results obtained in search and methodology. Conclusion:Although the Friedewald and Martin equations have a good correlation with low density lipoproteins as measured by direct methodology, studies that relate atherosclerotic arterial diseases to low density lipoproteins should consider the direct quantification of this lipoprotein in order to cover individuals with their different specificities.