Furoxan derivatives demonstrated in vivo efficacy by reducing Mycobacterium tuberculosis to undetectable levels in a mouse model of infection.

OBJECTIVES: The most recent survey conducted by the World Health Organization described Tuberculosis (TB) as one of the top 10 causes of death and the leading cause of death from a single infectious agent. The increasing number of TB-resistant cases has contributed to this scenario. In light of this, new strategies to control and treat the disease are necessary. Our research group has previously described furoxan derivatives as promising scaffolds to be explored as new antitubercular drugs. RESULTS: Two of these furoxan derivatives, (14b) and (14c), demonstrated a high selectivity against Mycobacterium tuberculosis. The compounds (14b) and (14c) were also active against a latent M. tuberculosis strain, with MIC90 values of 6.67 µM and 9.84 µM, respectively; they were also active against monoresistant strains (MIC90 values ranging from 0.61 to 20.42 µM) and clinical MDR strains (MIC90 values ranging from 3.09 to 42.95 µM). Time-kill experiments with compound (14c) showed early bactericidal effects that were superior to those of the first- and second-line anti-tuberculosis drugs currently used in therapy. The safety of compounds (14b) and (14c) was demonstrated by the Ames test because these molecules were not mutagenic under the tested conditions. Finally, we confirmed the safety, and high efficacy of compounds (14b) and (14c), which reduced M. tuberculosis to undetectable levels in a mouse aerosol model of infection. CONCLUSION: Altogether, we have identified two advanced lead compounds, (14b) and (14c), as novel promising candidates for the treatment of TB infection.

Byrsonima fagifolia Niedenzu Apolar Compounds with Antitubercular Activity.

Bioassay-guided fractionation of the chloroform extract of Byrsonima fagifolia leaves led to the isolation of active antitubercular compounds alkane dotriacontane (Minimal Inhibitory Concentration-MIC, 62.5 µg mL(-1)), triterpenoids as bassic acid (MIC = 2.5 µg mL(-1)), α-amyrin acetate (MIC = 62.5 µg mL(-1)), a mixture of lupeol, α- and ß-amyrin (MIC = 31.5 µg mL(-1)) and a mixture of lupeol, and acetates of α- and ß-amyrin (MIC = 31.5 µg mL(-1)). The antimycobacterial activity was determined by the Microplate Alamar Blue Assay (MABA) and the structures of promising compounds were determined by spectroscopic analysis. This investigation constitutes the first report of a chemical and antitubercular study of apolar compounds from B. fagifolia Niedenzu (IK).

Antimycobacterial flavones from Haplopappus sonorensis.

Crude extracts of Haplopappus sonorensis (A. Gray) S.F. Blake (Asteraceae), showed activity against Mycobacterium tuberculosis H(37)Rv. By assay-guided fractionation, 5-hydroxy-3,7,4'-trimethoxyflavone (1). 5,7-dihydroxy-3,4'-dimethoxyflavone (2). and 5,4'-dihydroxy-3,7-dimethoxyflavone (3). were identified as the antimycobacterial principles. Compound 2 was the most active compound.

Use of receiver operating characteristic curves to assess the performance of a microdilution assay for determination of drug susceptibility of clinical isolates of Mycobacterium tuberculosis.

The aim of this study was to apply receiver operating characteristic (ROC) analysis to the microplate Alamar blue assay, a recently developed alternative for drug susceptibility testing of mycobacteria. As this is a quantitative assay, its performance can be determined by ROC analysis, in which the area under the ROC curve represents a summary of test performance (the higher the area, the better the test's performance). Sixty isolates of Mycobacterium tuberculosis were tested by the microcolorimetric assay against six twofold dilutions of streptomycin, isoniazid, rifampin, and ethambutol. For each isolate, the susceptibility pattern was simultaneously established by the agar proportion method, the result of which represented the gold standard value for the ROC analysis. The critical concentration, area under the curve, and P value for each drug were determined by ROC curve analysis. The results of the assay were obtained in an average of 8 days of incubation. The performance of the assay was excellent for all four drugs: the area under the curves was >0.97, the P values were 0.000, and sensitivity was 94%, specificity 97%, predictive value for resistance >/=92%, predictive value for susceptibility 97%, and test efficiency 97%. According to ROC analysis, the microplate Alamar blue assay is a reliable method for determination of drug-susceptibility. Rapidity and cost efficiency are two additional qualities that make this test an excellent alternative for the drug susceptibility testing of Mycobacterium tuberculosis. The ROC curve analysis is a robust statistical approach for evaluating the performance of new quantitative methods for determination of drug sensitivity of Mycobacterium tuberculosis isolates.

Evaluation of the flora of Puerto Rico for in vitro antiplasmodial and antimycobacterial activities.

The emergence of resistant strains of Plasmodium falciparum and Mycobacterium tuberculosis underscores the need for novel drugs that are effective against these microorganisms. As part of our screening programme of the flora of Puerto Rico, we tested a number of ethanol extracts of higher plants for antiplasmodial and antimycobacterial activities. A total of 40 extracts belonging to 23 plant families and 37 species were tested for antiplasmodial activity. Five extracts demonstrated activity against Plasmodium falciparum in vitro (50%-100% parasite suppression at 5 microg/mL). Another 63 extracts belonging to 30 plant families and 50 species were tested in vitro against Mycobacterium tuberculosis. Two extracts were found to be active, Ficus citrifolia and Pisonia borinquena (85% or more inhibition of microbial growth at 100 microg/mL of extract).

Inhibition of Mycobacterium tuberculosis growth by saringosterol from Lessonia nigrescens.

Assay-guided fractionation of an antitubercular extract obtained from Lessonia nigrescens yielded the phytosterol saringosterol as its active component. No appreciable toxicity against Vero cells was observed for this compound. Saringosterol was also synthesized by oxidation of fucosterol. The MIC values for antitubercular activity of saringosterol and its 24S and 24R epimers were determined as 0.25, 1, and 0.125 microg/mL.

Antitubercular activity of triterpenoids from Lippia turbinata.

Assay-guided fractionation of the antitubercular MeOH-CH(2)Cl(2) extract obtained from Lippia turbinata led to the isolation of four novel triterpenoids-3beta,25-epoxy-3alpha,21alpha-dihydroxy-22beta-(3-methylbut-2-en-1-oyloxy)olean-12-ene-28-oic acid (1); 3beta,25-epoxy-3alpha,21alpha-dihydroxy-22beta-angeloyloxyolean-12-ene-28-oic acid (2); 3beta,25-epoxy-3alpha,21alpha-dihydroxy-22beta-tigloyloxyolean-12-ene-28-oic acid (3); and 3beta,25-epoxy-3alpha-hydroxy-22beta-(2-methylbutan-1-oyloxy)olean-12-ene-28-oic acid (4)-together with the known triterpenoids lantanilic acid (5), camaric acid (6), lantanolic acid (7), and rehmannic acid (8). The MIC values of 1-8 for growth inhibition of Mycobacterium tuberculosis were determined in the radiorespirometric BACTEC system.

Phytochemical, morphological, and biological investigations of propolis from Central Chile.

Propolis from central Chile was investigated for its plant origin by microscopical analysis of pollen grains and leaf fragments found in the sample. The pollen grains that appear with significant higher frequency in the sample corresponded to four native and two introduced species, whereas leaf fragments corresponded to four native species. Seventeen phenolic compounds that belong to the phenylpropane, benzaldehyde, dihydrobenzofuran, or benzopyran classes, were isolated from an organic extract that was found to have a moderate growth inhibitory activity against Mycobacterium avium, M. tuberculosis, and two strains of Staphylococcus aureus. The components responsible for activity were determined.

Antitubercular activity of pentacyclic triterpenoids from plants of Argentina and Chile.

Screening of plants from South America for antitubercular activity and subsequent assay-guided fractionation resulted in the isolation and characterization of several pentacyclic triterpenoids. The MIC values of 22 triterpenoids were determined using the radiorespiratory BACTEC assay and range from 8 microM to above 128 microM. The structure-activity relationships are discussed.

A new antitubercular mulinane diterpenoid from Azorella madreporica Clos.

Bioactivity-guided fractionation of the petroleum ether extract of Azorella madreporica Clos has led to the isolation of the novel, antitubercular mulinane diterpenoid 1. The structure has been elucidated on the basis of its 1D and 2D NMR spectra and by comparison with mulinolic acid 2 and a dehydration product 3 obtained from 1. The MIC of 1 for growth inhibition of the H37Rv strain of Mycobacterium tuberculosis was determined as 20 microg/mL. LC-MS and NMR have suggested the presence of this new compound in four other species of Azorella.

Rapid, low-technology MIC determination with clinical Mycobacterium tuberculosis isolates by using the microplate Alamar Blue assay.

A colorimetric, microplate-based Alamar Blue assay (MABA) method was used to determine the MICs of isoniazid (INH), rifampin, streptomycin (SM), and ethambutol (EMB) for 34 Peruvian Mycobacterium tuberculosis isolates (including both pansensitive and multidrug-resistant strains) and the H37Rv strain by using bacterial suspensions prepared directly from solid media. Results for all isolates were available within 8 days. Discordant results were observed on initial tests for 3 of 16 INH-susceptible isolates, 5 of 31 EMB-susceptible isolates, and 2 of 4 SM-resistant isolates (by the BACTEC 460 system). The overall agreements between the MICs obtained by MABA and the results obtained with the BACTEC 460 system were 87.9% for initial results and 93.6% after retesting 12 of 17 samples with discrepant results. Interpretation of MABA endpoints improved with technical experience. The MABA is a simple, rapid, low-cost, appropriate technology which does not require expensive instrumentation and which makes use of a nontoxic, temperature-stable reagent.