Determinação simultânea de Rifampicina, Isoniazida, Pirazinamida e Etambutol em comprimidos com doses fixas combinadas / Method for simultaneously determining the Rifampin, Isoniazid, Pyrazinamide and Ethambutol in the combined fixed-dose tablets

Os comprimidos utilizados no tratamento da tuberculose possuem quatro fármacos associados, isoniazida, pirazinamida, etambutol e rifampicina, e são distribuídos gratuitamente pelo Sistema Único de Saúde. Os métodos analíticos oficiais para analisar este medicamento estão especificados na Farmacopeia Americana 36a edição e na Farmacopeia Internacional 4a edição. Porém, estes compêndios oficiais não possuem monografias para análise simultânea dos quatro fármacos. O objetivo deste estudo foi desenvolver uma metodologia para determinar simultaneamente os princípios ativos em comprimidos dose fixa combinada, utilizando-se cromatografia a líquido de alta eficiência com detector de ultravioleta-visível, pois é de grande importância para o controle da qualidade do medicamento. O método desenvolvido utilizou coluna cromatográfica C18 (250 x 4,6) mm e 5 μm, fase móvel constituída de fase aquosa (85 % tampão formiato de amônio 0,3 mol/L pH 5, 15 % metanol e 0,005 mol/L de Cu2+ ou 250 mg/L de CuSO4.5H2O) e fase orgânica (metanol, 0,1 % de trietilamina e 0,2 % de ácido fórmico). O fluxo foi de 1,0 mL/min e comprimento de onda de 265 nm para isoniazida, pirazinamida e o etambutol e de 335 nm para rifampicina. Este método apresentou desvio padrão relativo inferior a 2,0 % na precisão e linearidade para os quatro fármacos estudados.

Tablets containing isoniazid, pyrazinamide, ethambutol and rifampicin are used for tuberculosis treatment, and theyare freely distributed by the Brazilian National Health System. The official analytical methods for testing those substances in fixed-dose combined tablet are described in the United States Pharmacopeia and theInternational Pharmacopoeia. None of these official compendiums refers to the methodologies forconducting the simultaneous analysis of these four drugs. This study aimedatdeveloping an analytical methodology to determine simultaneously allof four drugs in tablets for tuberculosis treatment, bymeans ofhigh performance liquid chromatography with ultraviolet-visible detector. The developed method used a chromatographic column with octadecylsilane stationary phase (250 mm x 4.6 mm x 5 μm particle size). The mobile phase was aqueous (85 % ammonium formate buffer pH 5, 15 % methanol and 250 mg CuSO4.5H2O), and organic phase (methanol, 0.1 % triethylamine and 0.2 % formic acid). The flow was 1.0 mL/min, at a wavelength of 265 nm or, when the equipment allowed, a wavelength of 265 nm for isoniazid, pyrazinamide and ethambutol and 335 nm for rifampicin. The developed methodology showed satisfactory results regarding the precision parameter, with relative standard deviation lower than 2.0 % for the studied drugs.

Resistencia a drogas antituberculosas en Caracas, Venezuela; 2001-2006 / Antituberculosis drug resistance in Caracas, Venezuela; 2001-2006

Para establecer la sensibilidad de las cepas de Mycobac-terium tuberculosis aisladas en Caracas, entre 2001 y 2006, fueron probadas utilizando el método colorimétrico para determinar las Concentraciones Inhibitorias Mínimas (CIM). De las 324 cepas, 46 (14,2%) mostraron resistencia a una o más drogas. Encontramos resistencia de alto nivel (CIM 8 µg/ml) y bajo nivel (CIM 1-4 µg/ml) a Estreptomicina en 6 (1,8%) y 25 (7,7%) de las cepas, respectivamente. Se encontró resistencia a Isoniacida de bajo nivel (MIC 0,125 - 0,5 µg/ml) en 8 (2,5%) y de alto nivel (MIC 1,0 µg/ml) en 15 (4,6%) de las cepas estudiadas. Hallamos 13 (4,0%) cepas resistentes a Rifampicina (RIF) (5 µg/ml) y 11 (3,4%) a Etambutol (10 µg/ml). De los 17 (5,2%) aislamientos resistentes a dos o más drogas, 12 (3,7%) fueron resistentes a INH y RIF (definido como multirresistencia, MDR). De las 12 cepas MDR, 11 fueron aisladas a partir de esputo y una de líquido pleural. Este estudio muestra un incremento en la prevalencia de la resistencia a las drogas antituberculosas en Caracas, especialmente las cepas MDR. Este aumento apunta hacia la necesidad de una encuesta na-cional, para evaluar el panorama real de la resistencia.

To asses drug susceptibility of Mycobacterium tuberculosis strains isolated from 2001 to 2006 in Caracas. Available strains were tested using colorimetric method to determine the Minimal Inhibitory Concentrations (MIC). Of 324 strains, 46 (14,2%) showed resistance to one or more drugs. High-resistance (8 µg/ml) and low-resistance (1-4 µg/ml) to Strep omycint was found in 6 (1,8%) and 25 (7,7%) strains, respectively. Isoniazid (INH) low-resistance (MIC 0.125 - 0.5 µg/ml) were found in 8 (2,5%) and high-resistant (MIC at 1.0 µg/ml) in 15 (4,6%), Rifampicin resistance (RMP) (5 µg/ml) in 13 (4%), and Ethambutol resistance (10 µg/ml) in 11 (3,4%) of the strains. Of the 17 (5,2%) isolates resistant to two or more drugs, 12 (3,7%) were resistant to INH and RMP (defined as multidrug resistance, MDR). Of these 12 MDR strains, 11 were isolated from sputum and one from pleu ral fluid. This study shows an increased prevalence of resistance to anti-tuberculosis drugs in Caracas, especially the prevalence of MDR strains, raises an urgent need of a proper nationwide survey to evaluate the true picture of resistance.

Concordancia de métodos para susceptibilidad antimicrobiana en cepas de Mycobacterium tuberculosis aisladas en Montería, Córdoba: tubo indicador de crecimiento micobacteriano vs. método de las proporciones múltiples / Agreement between methods for antimicrobial susceptibility to Mycobacterium tuberculosis strains isolated in Montería, Córdoba: mycobacteria growth indicator tube vs. proportion method

Objetivo: Evaluar la concordancia del método manual MGIT AST SIRE (mycobacteria growth indicator tube for antimicrobial susceptibility testing to isoniazid, rifampin, ethambutol and streptomycin) frente al método de proporciones múltiples (PM) para determinar susceptibilidad antimicrobiana en cepas de Mycobacterium tuberculosis. Métodos: Se analizaron 45 cepas de M. tuberculosis aisladas en la ciudad de Montería, entre 2003 y 2005, para susceptibilidad a (INH), rifampicina (RMP), etambutol (EMB) y estreptomicina (SM). El anßlisis de concordancia del método MGIT manual vs PM, se realizó con la prueba Kappa. Resultados: De las 45 cepas analizadas, 38 tuvieron idéntico resultado con ambos métodos y 7 tuvieron resultados discordantes. Los porcentajes globales de resistencia a drogas antituberculosas de las cepas analizadas por el MGIT manual y las PM fueron de 33.3% y 31.1%, respectivamente, la MDR fue 5 (11.1%) por PM y de 4 (8.8%) por MGIT manual. La concordancia del MGIT AST SIRE con el PM fue 95.5% para cada una de las drogas analizadas y kappa global de 0.8374. En cada antibiótico se encontraron dos cepas discordantes para INH, RIF (un falso resistente y un falso susceptible). En STR y EMB (dos falsos sensibles). El tiempo promedio de resultado para cualquiera de los antimicrobianos fue 6.5 días (rango 5-8 días) por MGIT, mientras que para PM fue 20 a 25 días. Conclusión: El presente estudio mostró que MGIT manual es concordante con PM, es rßpido, sencillo y eficaz, para determinar la susceptibilidad de cepas de M. tuberculosis.

Objective: To evaluate the agreement of manual MGIT (mycobacteria growth indicator tube for antimicrobial susceptibility testing to isoniazid, rifampin, ethambutol and streptomycin) for susceptibility testing vs proportion method on Lowenstein-Jensen (PM) in Mycobacterium tuberculosis strains. Methods: A total of forty-five isolates of M. tuberculosis were tested for susceptibility to isoniazid (INH), rifampin (RMP), ethambutol (EMB), and streptomycin (SM). The strains were isolated in Montería, during 2003 and 2005. The agreement between the two assays mentioned was estimated by the Kappa test. Results: There were thirty-eight strains with identical results while 7 had discrepant results with both methods. The overall resistance to antituberculosis drugs were 33.3% y 31.1% for manual MGIT and PM, respectively. MDR was 5 (11.1%) by PM and 4 (8.8%) by manual MGIT. The agreement between MGIT AST and PM was 95.5% for all drug tested and overall kappa value 0.8374. Two discrepancies were found in each drug; with INH and RIF (one false resistant and one false susceptible). STR and EMB (two false susceptibles). Turnaround times were 5 to 8 days (median, 6.5 days) for MGIT and 20 to 25 days for MP. Conclusions: These preliminary data show a good level of agreement between manual MGIT AST SIRE and MP. Also MGIT is a rapid, easy and efficient method for the drug susceptibility testing of M. tuberculosis.