Dihydrofolate Reductase Inhibitors: The Pharmacophore as a Guide for Co-Crystal Screening.

In this work, co-crystal screening was carried out for two important dihydrofolate reductase (DHFR) inhibitors, trimethoprim (TMP) and pyrimethamine (PMA), and for 2,4-diaminopyrimidine (DAP), which is the pharmacophore of these active pharmaceutical ingredients (API). The isomeric pyridinecarboxamides and two xanthines, theophylline (THEO) and caffeine (CAF), were used as co-formers in the same experimental conditions, in order to evaluate the potential for the pharmacophore to be used as a guide in the screening process. In silico co-crystal screening was carried out using BIOVIA COSMOquick and experimental screening was performed by mechanochemistry and supported by (solid + liquid) binary phase diagrams, infrared spectroscopy (FTIR) and X-ray powder diffraction (XRPD). The in silico prediction of low propensities for DAP, TMP and PMA to co-crystallize with pyridinecarboxamides was confirmed: a successful outcome was only observed for DAP + nicotinamide. Successful synthesis of multicomponent solid forms was achieved for all three target molecules with theophylline, with DAP co-crystals revealing a greater variety of stoichiometries. The crystalline structures of a (1:2) TMP:THEO co-crystal and of a (1:2:1) DAP:THEO:ethyl acetate solvate were solved. This work demonstrated the possible use of the pharmacophore of DHFR inhibitors as a guide for co-crystal screening, recognizing some similar trends in the outcome of association in the solid state and in the molecular aggregation in the co-crystals, characterized by the same supramolecular synthons.

Tuning down the environmental interests of organoclays for emerging pollutants: Pharmaceuticals in presence of electrolytes.

The impact of electrolytes on the adsorption of emerging pollutants: pharmaceuticals onto layered materials: a raw clay mineral and its nonionic and cationic organoclay derivatives was studied. The selected pharmaceuticals: amoxicillin, norfloxacin, sulfamethoxazole, metoprolol, carbamazepine, and trimethoprim show different electric charges: zwitterionic, anionic, cationic and neutral and hydrophobic character (different LogP). Without any salts, the set of complementary data obtained by UV and infrared spectroscopies, X-ray diffraction points out the importance of the electric charge which represents a key parameter in both the spontaneity and feasibility of the adsorption. In contrast, the hydrophobicity of the analytes plays a minor role but determines the magnitude of the adsorbed amount of pharmaceuticals onto organoclays. With a dual hydrophilic and hydrophobic behavior, nonionic organoclay appears to be the most polyvalent material for the removal of the pharmaceuticals. In the presence of electrolytes (NaCl at a concentration of 1 × 10-2 mol L-1), both nonionic and cationic organoclays show a decrease of their efficiencies, whereas the adsorption is particularly enhanced for Na-Mt except for the cationic species (trimethoprim and metoprolol). Thus, in realistic experimental conditions close to those of natural effluents, raw clay mineral appears as the most appropriate sorbent for the studied pharmaceuticals while it raises the question of the usefulness of organoclays in water remediation strategy.

Effects of Microplate Type and Broth Additives on Microdilution MIC Susceptibility Assays.

The determination of antibiotic potency against bacterial strains by assessment of their minimum inhibitory concentration normally uses a standardized broth microdilution assay procedure developed more than 50 years ago. However, certain antibiotics require modified assay conditions in order to observe optimal activity. For example, daptomycin requires medium supplemented with Ca2+, and the lipoglycopeptides dalbavancin and oritavancin require Tween 80 to be added to the growth medium to prevent the depletion of free drug via adsorption to the plastic microplate. In this report, we examine systematically the effects of several different plate types on microdilution broth MIC values for a set of antibiotics against Gram-positive and Gram-negative bacteria, both in medium alone and in medium supplemented with the commonly used additives Tween 80, lysed horse blood, and 50% human serum. We observed very significant differences in measured MICs (up to 100-fold) for some lipophilic antibiotics, such as the Gram-positive lipoglycopeptide dalbavancin and the Gram-negative lipopeptide polymyxins, and found that nonspecific binding plates can replace the need for surfactant additives. Microtiter plate types and any additives should be specified when reporting broth dilution MIC values, as results can vary dramatically for some classes of antibiotics.

NMR structures of apo L. casei dihydrofolate reductase and its complexes with trimethoprim and NADPH: contributions to positive cooperative binding from ligand-induced refolding, conformational changes, and interligand hydrophobic interactions.

In order to examine the origins of the large positive cooperativity (ΔG(0)(coop) = -2.9 kcal mol(-1)) of trimethoprim (TMP) binding to a bacterial dihydrofolate reductase (DHFR) in the presence of NADPH, we have determined and compared NMR solution structures of L. casei apo DHFR and its binary and ternary complexes with TMP and NADPH and made complementary thermodynamic measurements. The DHFR structures are generally very similar except for the A-B loop region and part of helix B (residues 15-31) which could not be directly detected for L. casei apo DHFR because of line broadening from exchange between folded and unfolded forms. Thermodynamic and NMR measurements suggested that a significant contribution to the cooperativity comes from refolding of apo DHFR on binding the first ligand (up to -0.95 kcals mol(-1) if 80% of A-B loop requires refolding). Comparisons of Cα-Cα distance differences and domain rotation angles between apo DHFR and its complexes indicated that generally similar conformational changes involving domain movements accompany formation of the binary complexes with either TMP or NADPH and that the binary structures are approaching that of the ternary complex as would be expected for positive cooperativity. These favorable ligand-induced structural changes upon binding the first ligand will also contribute significantly to the cooperative binding. A further substantial contribution to cooperative binding results from the proximity of the bound ligands in the ternary complex: this reduces the solvent accessible area of the ligand and provides a favorable entropic hydrophobic contribution (up to -1.4 kcal mol(-1)).

Oxidation of antibiotics during water treatment with potassium permanganate: reaction pathways and deactivation.

Recent work demonstrates that three widely administered antibiotics (ciprofloxacin, lincomycin, and trimethoprim) are oxidized by potassium permanganate [KMnO(4), Mn(VII)] under conditions relevant to water treatment operations. However, tests show that little to no mineralization occurs during reactions with Mn(VII), so studies were undertaken to characterize the reaction products and pathways and to assess the effects of Mn(VII)-mediated transformations on the antibacterial activity of solutions. Several oxidation products were identified for each antibiotic by liquid chromatography-tandem mass spectrometry (LC-MS/MS). For ciprofloxacin, 12 products were identified, consistent with oxidation of the tertiary aromatic and secondary aliphatic amine groups on the piperazine ring and the cyclopropyl group. For lincomycin, seven products were identified that indicate structural changes to the pyrrolidine ring and thioether group. For trimethoprim, seven products were identified, consistent with Mn(VII) reaction at C═C double bonds on the pyrimidine ring and the bridging methylene group. Oxidation pathways are proposed based on the identified products. Bacterial growth inhibition bioassays (E. coli DH5α) show that the mixture of products resulting from Mn(VII) reactions with the antibiotics collectively retain negligible antibacterial potency in comparison to the parent antibiotics. These results suggest that permanganate can be an effective reagent for eliminating the pharmaceutical activity of selected micropollutants during drinking water treatment.

Oxidation kinetics of antibiotics during water treatment with potassium permanganate.

The ubiquitous occurrence of antibiotics in aquatic environments raises concerns about potential adverse effects on aquatic ecology and human health, including the promotion of increased antibiotic resistance. This study examined the oxidation of three widely detected antibiotics (ciprofloxacin, lincomycin, and trimethoprim) by potassium permanganate [KMnO(4); Mn(VII)]. Reaction kinetics were described by second-order rate laws, with apparent second-order rate constants (k(2)) at pH 7 and 25 degrees C in the order of 0.61 +/- 0.02 M(-1) s(-1) (ciprofloxacin) < 1.6 +/- 0.1 M(-1) s(-1) (trimethoprim) < 3.6 +/- 0.1 M(-1) s(-1) (lincomycin). Arrhenius temperature dependence was observed with apparent activation energies (E(a)) ranging from 49 kJ mol(-1) (trimethoprim) to 68 kJ mol(-1) (lincomycin). Rates of lincomycin and trimethoprim oxidation exhibited marked pH dependences, whereas pH had only a small effect on rates of ciprofloxacin oxidation. The effects of pH were quantitatively described by considering parallel reactions between KMnO(4) and individual acid-base species of the target antibiotics. Predictions from a kinetic model that included temperature, KMnO(4) dosage, pH, and source water oxidant demand as input parameters agreed reasonably well with measurements of trimethoprim and lincomycin oxidation in six drinking water utility sources. Although Mn(VII) reactivity with the antibiotics was lower than that reported for ozone and free chlorine, its high selectivity and stability suggests a promising oxidant for treating sensitive micropollutants in organic-rich matrices (e.g., wastewater).

Evaluating the vulnerability of surface waters to antibiotic contamination from varying wastewater treatment plant discharges.

Effluents from three wastewater treatment plants with varying wastewater treatment technologies and design were analyzed for six antibiotics and caffeine on three sampling occasions. Sulfamethoxazole, trimethoprim, ciprofloxacin, tetracycline, and clindamycin were detected in the effluents at concentrations ranging from 0.090 to 6.0 microg/L. Caffeine was detected in all effluents at concentrations ranging from 0.19 to 9.9 microg/L. These findings indicate that several conventional wastewater management practices are not effective in the complete removal of antibiotics, and their discharges have a large potential to affect the aquatic environment. To evaluate the persistence of antibiotics coming from the wastewater discharges on the surrounding surface waters, samples were collected from the receiving streams at 10-, 20- and 100-m intervals. Ciprofloxacin, sulfamethoxazole, and clindamycin (0.043 to 0.076 microg/L) were found as far as 100 m from the discharge point, which indicates the persistence of these drugs in surface waters.

[Preliminary study on isoniazid-epiroprim combination in a tuberculosis murine model]. / Etude préliminaire de l'association isoniazide-épiroprim dans un modèle murin de tuberculose.

The purpose of this study regarding isoniazid-epiroprim's association applied to antituberculosis chemotherapy, carried through murine model, initiated into Institut Pasteur of Côte d'Ivoire and worked out at Institut Pasteur of Paris was to evaluate the epiroprim's effect alone and associated with isoniazid on Mycobacterium tuberculosis. Sixteen mouses (lineage C57Bl/6) were inoculated by venous way with 10(5) viable bacillus (strain H37Rv) suspended in 500 microliters sterile physiological aqueous solution and were shared out into 4 sets. Fifteen days later the sets have been submitted or not to a daily treatment by gavage during three weeks (epiroprim, isoniazid, isoniazid plus epiroprim). The mouses were euthanasied, spleen and lung were removed from each animal. The titres of determined bacillus into those organs prove that isoniazid and epiroprim associated seem more efficacious than the isoniazid monotherapy for mouses pulmonary tuberculosis. Bacillus obtained are sensitive to isoniazid.

Automated analytical systems for drug development studies. II--A system for dissolution testing.

Microdialysis is a non-equilibrium dynamic sampling method in which the analytes diffuse across a semipermeable membrane due to a concentration gradient and are carried away by the constantly pumping perfusion medium for on-line analysis. A BAS, Inc. microinfusion pump/injector and an on-line LC system were interfaced with a dissolution apparatus to automate dissolution testing of tablets. A DL-5 microdialysis loop probe was suspended in the dissolution medium for sampling. The automated system was used reproducibly (RSD < 2%) to measure the dissolution of acetaminophen and Sulfatrim tablets. Drug recovery from the microdialysis probe was a function of the perfusion rate at constant temperature. However, microdialysis recovery was independent of drug concentration over the linear ranges of the assays for the analytes of interest. Dissolution profiles determined by microdialysis sampling were compared with manual sampling. Identical profiles were obtained for acetaminophen tablets in water at 37 degrees C and 50 rpm by both sampling methods. Dissolution of Sulfatrim tablets was determined in 0.1 M hydrochloride acid at 37 degrees C and 75 rpm. Microdialysis sampling permitted the use of a specially designed perfusion medium to buffer the acidic samples before injecting onto the LC column. Dissolution profiles of sulphamethoxazole were comparable for both sampling methods; however, microdialysis sampling indicated slightly higher release of trimethoprim from the Sulfatrim tablets, which was attributed to release of adsorbed drug from the connecting tubing.