Structure, Solubility and Stability of Orbifloxacin Crystal Forms: Hemihydrate versus Anhydrate.

Orbifloxacin (ORBI) is a widely used antimicrobial drug of the fluoroquinolone class. In the official pharmaceutical compendia the existence of polymorphism in this active pharmaceutical ingredient (API) is reported. No crystal structure has been reported for this API and as described in the literature, its solubility is very controversial. Considering that different solid forms of the same API may have different physicochemical properties, these different solubilities may have resulted from analyses inadvertently carried out on different polymorphs. The solubility is the most critical property because it can affect the bioavailability and may compromise the quality of a drug product. The crystalline structure of ORBI determined by SCXRD is reported here for the first time. The structural analysis reveals that the ORBI molecule is zwitterionic and hemihydrated. ORBI hemihydrated form was characterized by the following techniques: TG/DTA, FTIR-ATR, and PXRD. A second crystalline ORBI form is also reported: the ORBI anhydrous form was obtained by heating the hemihydrate. These ORBI solid forms were isomorphous, since no significant change in unit cell and space group symmetry were observed. The solid-state phase transformation between these forms is discussed and the equilibrium solubility data were examined in order to check the impact of the differences observed in their crystalline structures.

Synthesis and biological evaluation against Leishmania amazonensis of a series of alkyl-substituted benzophenones.

Nine O-alkyl and O-prenyl derivatives were synthesized from commercial 2,4-dihydroxybenzophenone, 4,e4,4'-dihydroxybenzophenone and were evaluated for their leishmanicidal activity against promastigote forms of Leishmania amazonensis, as well their toxicity in murine macrophages. All derivatives exhibited better biological activity than their hydroxylated benzophenones precursors, and new compound LFQM-123 (3c) was 250-fold more active than its precursor 4,4'-dihydroxybenzophenone (3). Moreover, some of the results were comparable to the standard drug Amphotericin B, suggesting that the increase in lipophilicity could facilitate protozoa membrane permeation. In this study we confirmed that benzophenone derivatives exhibit leishmanicidal properties, with relatively low toxicity, and thus could be exploited as promise prototypes for the design and development of new drug for the treatment of leishmaniasis.

Inhibition of cysteine proteases by a natural biflavone: behavioral evaluation of fukugetin as papain and cruzain inhibitor.

Cruzain is the major cysteine protease of Trypanosoma cruzi, the infectious agent responsible for Chagas disease, and cruzain inhibitors display considerable antitrypanosomal activity. In the present work we elucidated crystallographic data of fukugetin, a biflavone isolated from Garcinia brasiliensis, and investigated the role of this molecule as cysteine protease inhibitor. The kinetic analyses demonstrated that fukugetin inhibited cruzain and papain by a slow reversible type inhibition with K(I) of 1.1 and 13.4 µM, respectively. However, cruzain inhibition was about 12 times faster than papain inhibition. Lineweaver-Burk plots demonstrated partial competitive inhibition for cruzain and hyperbolic mixed-type inhibition for papain. Furthermore, the docking results showed that the biflavone binds to ring C' in the S2 pocket and to ring C in the S3 pocket through hydrophobic interactions and hydrogen bonds. Finally, fukugetin also presented inhibitory activity on proteases of the T. cruzi extract, with IC50 of 7 µM.

Synthesis, characterization and in vitro cytotoxicity of ruthenium(II) metronidazole complexes: Cell cycle arrest at G1/S transition and apoptosis induction in MCF-7 cells.

Ruthenium compounds are known to be potential drug candidates since they offer the potential for reduced toxicity. Furthermore, the various oxidation states, different mechanisms of action and ligand substitution kinetics give them advantages over platinum-based complexes, making them suitable for use in biological applications. So, herein, novel ruthenium(II) complexes with metronidazole as ligand were obtained [RuCl(MTNZ)(dppb)(4,4'-Mebipy)]PF6 (1), [RuCl(MTNZ)(dppb)(4,4'-Methoxybipy)]PF6 (2), [RuCl(MTNZ)(dppb)(bipy)]PF6 (3) and [RuCl(MTNZ)(dppb)(phen)]PF6 (4) where, MTNZ = metronidazole, dppb = 1,4-bis(diphenylphosphino)butane, 4,4'-Mebipy = 4,4'-dimethyl-2,2'-bipyridine, 4,4'-Methoxybipy = 4,4'-dimethoxy-2,2'-bipyridine, bipy = 2,2'-bipyridine and phen = 1,10-phenanthroline. The complexes were characterized by elemental analysis, molar conductivity, infrared and UV-Vis spectroscopy, cyclic voltammetry, 31P{1H}, 1H, 13C{1H} and Dept 135 NMR and mass spectrometry. The interaction of complexes 1-4 with DNA was evaluated, and their cytotoxicity profiles were determined on four different tumor cell lines derived from human cancers (SK-MEL-147, melanoma; HepG2, hepatocarcinoma; MCF-7, estrogen-positive breast cancer; A549, non-small cell lung cancer). We demonstrated that complexes (1) and (3) are promising antitumor agents once inhibited the proliferative behavior of MCF-7 cells and induced apoptosis.

Assessment of the biological potential of diaryltriazene-derived triazene compounds.

In the present study, novel, 1,3-diaryltriazene-derived triazene compounds were synthesized and tested. Triazenes are versatile and belong to a group of alkylating agents with interesting physicochemical properties and proven biological activities. This study describes the synthesis, molecular and crystalline structure, biological activity evaluation, and antifungal and antimicrobial potentials of 1,3-bis(X-methoxy-Y-nitrophenyl)triazenes [X = 2 and 5; Y = 4 and 5]. The antimicrobial and antifungal activities of the compounds were tested by evaluating the sensitivity of bacteria (American Type Culture Collection, ATCC) and clinical isolates to their solutions using standardized microbiological assays, cytotoxicity evaluation, and ecotoxicity tests. The antimicrobial potentials of triazenes were determined according to their minimum inhibitory concentrations (MICs); these compounds were active against gram-positive and gram-negative bacteria, with low MIC values. The most surprising result was obtained for T3 having the effective MIC of 9.937 µg/mL and antifungal activity against Candida albicans ATCC 90028, C. parapsilosis ATCC 22019, and C. tropicallis IC. To the best of our knowledge, this study is the first to report promising activities of triazene compounds against yeast and filamentous fungi. The results showed the potential utility of triazenes as agents affecting selected resistant bacterial and fungal strains.

Buclizine crystal forms: First Structural Determinations, counter-ion stoichiometry, hydration, and physicochemical properties of pharmaceutical relevance.

Buclizine (BCZ) is a chiral synthetic piperazine derivative which has antihistaminic, anti-muscarinic and antiemetic properties, and has been reintroduced as an appetite stimulant, especially for pediatric patients. Structural information about this drug, as well as other buclizine crystalline forms (solvates, salts and co-crystals) including the BCZ free-base (BCZ-FB), is non-existent. Here, we present for the first time the crystal structure of the monohydrochloride monohydrate salt of BCZ (BCZHCl·H2O), and of its anhydrous form, BCZHCl. Interestingly, BCZHCl·H2O was obtained by recrystallization from the raw material (BCZH2Cl2) in ethanol:water solution showing that BCZ anhydrous dihydrochloride salt changes easily to a monohydrochloride monohydrate salt modification, which raise concerns about formulation quality control. BCZHCl·H2O and BCZHCl crystallize in the orthorhombic space groups (Pna21 and Pca21) belonging to the mm2 point group and are thus classified as non-centrosymmetric achiral structures (NA). Intuitively, we expect these salts to crystallize in a space group with a center of symmetry, since less than 5% of the known racemic compounds crystallize in the NA type. The crystal structures of BCZH2Cl2 and BCZ-FB were not determined, but their existence was verified by other techniques (chloride ion analysis, PXRD, HPLC, FT-IR, DSC, TGA) and by comparison of the obtained results with those found for BCZHCl. Additionally, we have also performed an evaluation of the equilibrium solubility (at six different aqueous media) and the dissolution profile of the BCZHCl salt compared to the raw material and BCZ-FB. Different equilibrium solubility values were found comparing the three forms in acidic and neutral pH ranges and all of them were insoluble at pH > 7.0. Moreover, tablets prepared with BCZH2Cl2, BCZHCl or BCZ-FB show significant differences in terms of dissolution profile.

Synthesis, crystal structure and leishmanicidal activity of new trimethoprim Ru(III), Cu(II) and Pt(II) metal complexes.

Leishmaniasis is a parasitic disease caused by protozoa of the genus Leishmania, which has very limited treatment options and affects poor and underdeveloped populations. The current treatment is plagued by many complications, such as high toxicity, high cost and resistance to parasites; therefore, novel therapeutic agents are urgently needed. Herein, the synthesis, characterization and in vitro leishmanicidal potential of new complexes with the general formula [RuCl3(TMP)(dppb)] (1), [PtCl(TMP)(PPh3)2]PF6 (2) and [Cu(CH3COO)2(TMP)2]·DMF (3) (dppb = 1,4-bis(diphenylphosphino)butane, PPH3 = triphenylphosphine and TMP = trimethoprim) were evaluated. The complexes were characterized by infrared, UV-vis, cyclic voltammetry, molar conductance measurements, elemental analysis and NMR experiments. Also, the geometry of (2) and (3) were determined by single crystal X-ray diffraction. Despite being less potent against promastigote L. amazonensis proliferation than amphotericin B reference drug (IC50 = 0.09 ± 0.02 µM), complex (2) (IC50 = 3.6 ± 1.5 µM) was several times less cytotoxic (CC50 = 17.8 µM, SI = 4.9) in comparison with amphotericin B (CC50 = 3.3 µM, SI = 36.6) and gentian violet control (CC50 = 0.8 µM). Additionally, complex (2) inhibited J774 macrophage infection and amastigote number by macrophages (IC50 = 6.6 and SI = 2.7). Outstandingly, complex (2) was shown to be a promising candidate for a new leishmanicidal therapeutic agent, considering its biological power combined with low toxicity.

Solid-State Characterization of Spironolactone 1/3 Hydrate.

Spironolactone (SPR) is a poorly water-soluble drug widely used for the treatment of various diseases. The objective of this study was to carry out the preparation and solid-state characterization of SPR 1/3 hydrate. The solid form was generated by an unreported recrystallization process in acetone and characterized for the first time by a combination of X-ray powder diffraction (XRD), differential scanning calorimetry (DSC), thermogravimetric analysis (TGA), Fourier-transform infrared spectroscopy (FTIR), equilibrium solubility, and an accelerated stability study. XRD, DSC, and TGA studies revealed that SPR 1/3 hydrated converts completely to form II after heating to 180°C. Solubility studies at 37°C showed that SPR 1/3 hydrate was statistically less soluble than SPR form II in all tested media and that SPR form II partially converts to SPR 1/3 hydrate in aqueous media. Accelerated stability studies demonstrated that both forms were physically and chemically stable up to 6 months (40°C/75% RH). We concluded that contamination of SPR 1/3 hydrate in SPR raw materials is undesirable. Taking this into account we recommend its polymorphic monitoring either in active pharmaceutical ingredients or commercial tablets by solid-state identification/quantification methods (XRD, DSC, TGA, and FTIR). Of these, XRD proved to be the most conclusive and accurate.

A novel platinum complex containing a piplartine derivative exhibits enhanced cytotoxicity, causes oxidative stress and triggers apoptotic cell death by ERK/p38 pathway in human acute promyelocytic leukemia HL-60 cells.

Piplartine (piperlongumine) is a plant-derived compound found in some Piper species that became a novel potential antineoplastic agent. In the present study, we synthesized a novel platinum complex containing a piplartine derivative cis-[PtCl(PIP-OH)(PPh3)2]PF6 (where, PIP-OH = piplartine demethylated derivative; and PPh3 = triphenylphosphine) with enhanced cytotoxicity in different cancer cells, and investigated its apoptotic action in human promyelocytic leukemia HL-60 cells. The structure of PIP-OH ligand was characterized by X-ray crystallographic analysis and the resulting platinum complex was characterized by infrared, molar conductance measurements, elemental analysis and NMR experiments. We found that the complex is more potent than piplartine in a panel of cancer cell lines. Apoptotic cell morphology, increased internucleosomal DNA fragmentation, without cell membrane permeability, loss of the mitochondrial transmembrane potential, increased phosphatidylserine externalization and caspase-3 activation were observed in complex-treated HL-60 cells. Treatment with the complex also caused a marked increase in the production of reactive oxygen species (ROS), and the pretreatment with N-acetyl-L-cysteine, an antioxidant, reduced the complex-induced apoptosis, indicating activation of ROS-mediated apoptosis pathway. Important, pretreatment with a p38 MAPK inhibitor (PD 169316) and MEK inhibitor (U-0126), known to inhibit ERK1/2 activation, also prevented the complex-induced apoptosis. The complex did not induce DNA intercalation in cell-free DNA assays. In conclusion, the complex exhibits more potent cytotoxicity than piplartine in a panel of different cancer cells and triggers ROS/ERK/p38-mediated apoptosis in HL-60 cells.

The foliar application of a mixture of semisynthetic chitosan derivatives induces tolerance to water deficit in maize, improving the antioxidant system and increasing photosynthesis and grain yield.

Research has shown that chitosan induces plant stress tolerance and protection, but few studies have explored chemical modifications of chitosan and their effects on plants under water stress. Chitosan and its derivatives were applied (isolated or in mixture) to maize hybrids sensitive to water deficit under greenhouse conditions through foliar spraying at the pre-flowering stage. After the application, water deficit was induced for 15 days. Analyses of leaves and biochemical gas exchange in the ear leaf were performed on the first and fifteenth days of the stress period. Production attributes were also analysed at the end of the experiment. In general, the application of the two chitosan derivatives or their mixture potentiated the activities of the antioxidant enzymes superoxide dismutase, catalase, ascorbate peroxidase, glutathione reductase and guaiacol peroxidase at the beginning of the stress period, in addition to reducing lipid peroxidation (malonaldehyde content) and increasing gas exchange and proline contents at the end of the stress period. The derivatives also increased the content of phenolic compounds and the activity of enzymes involved in their production (phenylalanine ammonia lyase and tyrosine ammonia lyase). Dehydroascorbate reductase and compounds such as total soluble sugars, total amino acids, starch, grain yield and harvest index increased for both the derivatives and chitosan. However, the mixture of derivatives was the treatment that led to the higher increase in grain yield and harvest index compared to the other treatments. The application of semisynthetic molecules derived from chitosan yielded greater leaf gas exchange and a higher incidence of the biochemical conditions that relieve plant stress.

Pro-apoptotic activity of ruthenium 1-methylimidazole complex on non-small cell lung cancer.

Herein, novel ruthenium(II) complexes containing 1-methylimidazole as a ligand were obtained with the following formulas: [RuCl(1Meim)(dppb)(bpy)]Cl (1), [RuCl(1Meim)(dppb)(4,4'-DMbpy)]Cl (2), [RuCl(1Meim)(dppb)(5,5'-DMbpy)]Cl (3) and [RuCl(1Meim)(dppb)(phen)]Cl (4) where, 1Meim = 1-methylimidazole, dppb = 1,4-Bis(diphenylphosphino)butane, bpy = 2,2'-bipyridine, 4,4'-DMbpy = 4,4'-dimethyl-2,2'-bipyridine, 5,5'-DMbpy = 5,5'-dimethyl-2,2'-bipyridine and phen = 1,10-phenanthroline. Additionally, crystal structures containing the cations of (1) and (3) were obtained when the counter ion was exchanged, leading to the formation of [RuCl(1Meim)(dppb)(bpy)]PF6 (5) and [RuCl(1Meim)(dppb)(5,5'-DMbpy)]PF6 methanol solvate (6) where PF6 = hexafluorophosphate, showing one 1-methylimidazole molecule coordinated through the imidazole nitrogen, as expected. The complexes were characterized by elemental analysis, molar conductivity, infrared and UV-Vis spectroscopy, 1H, 13C{1H} and 31P{1H} NMR, mass spectrometry and cyclic voltammetry. The interactions of complexes 1-4 with DNA and human serum albumin (HSA) were evaluated, and the cytotoxicity profiles of compounds 1-4 were determined using four different tumor cell lines derived from human cancers (melanoma: HT-144, colon: HCT-8, breast: MDA-MB-231 and lung: A549). A higher cytotoxic activity was observed for compound (3) against non-small cell lung cancer (A549). Complex (3) inhibited the clonogenic capacity and cell cycle progression of A549 cells and induced apoptosis involving mitochondrial pathway activation. Therefore, the data obtained in the present study support further investigations concerning molecular targets of complex (3) in non-small cell lung cancer.

Pharmacokinetics and pharmacodynamics of glimepiride polymorphs.

Glimepiride (GLIM) is used as an oral antihyperglycemic agent for treatment of type 2 diabetes. The drug presents two polymorphic forms (GLIM form I and GLIM form II) described in the literature, and according to in vitro data, GLIM form II is about 3.5 times more soluble and releases 2 times the drug amount than GLIM form I in the physiological pH range. Considering the literature in vitro data and that the diabetes treatment demands glycemic control, avoiding abrupt fluctuations in the blood glucose levels, this work aimed to study the impact of GLIM polymorphism in the in vivo performance of GLIM solid oral dosages. For this, hard gelatin capsules with GLIM form I or II were prepared and orally administered in rats. After that, pharmacokinetic studies were performed by sampling animal blood at different times, and biochemical parameters (pharmacodynamic), such as glucose and insulin, were also evaluated. Our results showed that the in vitro data corroborate with our in vivo assays. GLIM form II provided higher plasma concentration of drug than form I (at baseline up to approximately 200 min after oral administration) and, consequently, increased insulin release and reduced levels of glucose, showing good correlation between pharmacokinetic and pharmacodynamics assays. Thus, this study demonstrated that GLIM polymorphs in oral dosages might alter the drug efficacy, which may expose the patients to risks, such as hypoglycemia.

Analysis of polymorphic contamination in meloxicam raw materials and its effects on the physicochemical quality of drug product.

This work aims to evaluate the effect of polymorphism on the physicochemical properties of meloxicam, which is an antipyretic and non-steroidal anti-inflammatory drug. Powder X-ray Diffraction, Infrared Spectroscopy with attenuated total reflectance, Thermogravimetric and Differential Scanning Calorimetry techniques were used for the polymorphic characterization. Comparative tests of solubility, intrinsic dissolution and dissolution profiles were performed on meloxicam active pharmaceutical ingredients (APIs) and formulated tablets. A polymorphic contamination (Forms I and III) was found in a studied meloxicam batch, which showed a higher solubility and greater intrinsic dissolution than those containing only the preconized form (Form I). Consequently, the dissolution profiles of the tablets that contained the polymorphic contamination showed higher drug release. Additionally, a thermal behavior study shows that MLX Form I and III are monotropy polymorphs being MLX Form III a metastable phase, which becomes MLX Form I at approximately 200°C in solid state phase transition governed by kinetic variables. The kinetic of conversion of Form III to Form I in saturated solutions was also studied. These results illustrate the importance of the polymorphic characterization of meloxicam APIs and formulated tablets in order to avoid potential quality and efficacy problems of drug products.

Non-centrosymmetric crystals of new N-benzylideneaniline derivatives as potential materials for non-linear optics.

Three new N-benzylideneaniline derivatives [p-nitrobenzylidene-p-phenylamineaniline (I), 2,4-dinitrobenzylidene-p-phenylamineaniline (II) and p-dinitrobenzylidene-p-diethylamineaniline (III)] containing electron-push-pull groups have been prepared. They present a planar N-benzylideneaniline core and neighbouring functional atoms, which are related through an efficient intramolecular charge transfer (CT). Two of the derivatives crystallize in non-centrosymmetric space groups, a necessary condition for non-linear optical (NLO) responses. The NLO properties were calculated for the molecular conformations determined by single-crystal X-ray diffraction as well as for the four molecules packed into each corresponding unit cell, using a quantum-chemical method at the cam-B3LYP/NLO-V level of theory. As expected from antiparallel face-to-face stacking through centrosymmetry, the main NLO descriptors - namely, the first hyperpolarizability (ßtot) and its projection on the dipole moment direction (ßvec) - are almost zero for the tetramer of derivative III. Interestingly, the calculated first hyperpolarizability decreases in the non-centrosymmetric unit-cell content of derivative II when compared to its single molecule, which may be related to its molecular pillaring, similar to that observed in derivative III. On the other hand, a desirable magnification of the NLO properties was found for packed units of derivative I, which may be a consequence of its parallel face-to-tail stacking with the CT vectors of all molecules pointing in the same direction. Moreover, the CT vector of compound I makes an angle of θ = 33.6° with its crystal polar axis, resulting in a higher-order parameter (cos(3)θ = 0.6) compared with the other derivatives. This is in line with the higher macroscopic second-order NLO response predicted for derivative I, ßtot = 120.4 × 10(-30) e.s.u.

Analgesic and anti-inflammatory activities of the 2,8-dihydroxy-1,6-dimethoxyxanthone from Haploclathra paniculata (Mart) Benth (Guttiferae).

In the present study, the pharmacological effects of 2,8-dihydroxy-1,6-dimethoxyxanthone from the bark of Haploclathra paniculata were investigated in mice using in vivo inflammation and nociception models. Acetic acid-induced writhing, paw licking induced by formalin, hot plate, and carrageenan-induced paw edema tests were used to investigate the anti-inflammatory and antinociceptive activities of the xanthone compound. Xanthone, at both doses, inhibited abdominal writhing and the formalin test. At a dose of 20 mg/kg, the time of reaction to the hot plate increased, and significant effects were observed after 30, 60 and 90 min of treatment. At doses of 10 and 20 mg/kg p.o., the 2,8-dihydroxy-1,6-dimethoxyxanthone significantly reduced paw edema at 3 h after the stimulus. The tests also showed no acute toxicity of the xanthone compound in mice. 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging ability was also studied and confirmed the antioxidant activity of the xanthone. To propose the mechanism of action of anti-inflammatory activity of the xanthone, a molecular docking was performed using the isoenzymes cyclooxygenase 1 and 2 and the results indicate that the molecule is capable of inhibiting both the enzymes. Therefore, it can be concluded that 2,8-dihydroxy-1,6-dimethoxyxanthone from H. paniculata demonstrates analgesic, anti-inflammatory, and antioxidant activities.

Solubility and dissolution studies of tibolone polymorphs

ABSTRACT Different solid forms of an active pharmaceutical ingredient can have distinct chemical and physical characteristics. In this work, we studied the solubility and dissolution properties of the described tibolone polymorphic forms (I and II). Both forms were successively recrystallized and characterized by powder X-ray diffraction and attenuated total reflection infrared spectroscopy. Equilibrium solubility and dissolution profiles were performed for both forms. Solubility studies demonstrated that form II is statistically more soluble in water, 0.01 mol L-1 HCl and pH 4.5 acetate buffer. The solubility of forms I and II were explained in terms of crystal packing. Dissolution tests of tablets showed a lower release of polymorphic form II than form I from tablets. The results showed an impact of polymorphism on the quality of tibolone tablets and suggest that tibolone forms I and II can show distinct interactions with pharmaceutical excipients used in tablets. Therefore, only form I is acceptable for the preparation of tablet forms. Based on our results, we propose the quality control on tibolone raw materials using X-ray diffraction analysis and attenuated total reflection infrared spectroscopy.

Polymorphism: an evaluation of the potential risk to the quality of drug products from the Farmácia Popular Rede Própria

Polymorphism in solids is a common phenomenon in drugs, which can lead to compromised quality due to changes in their physicochemical properties, particularly solubility, and, therefore, reduce bioavailability. Herein, a bibliographic survey was performed based on key issues and studies related to polymorphism in active pharmaceutical ingredient (APIs) present in medications from the Farmácia Popular Rede Própria. Polymorphism must be controlled to prevent possible ineffective therapy and/or improper dosage. Few mandatory tests for the identification and control of polymorphism in medications are currently available, which can result in serious public health concerns.

O polimorfismo em sólidos é um fenômeno frequente em fármacos e pode levar a problemas na qualidade dos medicamentos por alterar suas propriedades físico-químicas, em especial a solubilidade e, consequentemente, a biodisponibilidade. Nesse trabalho realizou-se levantamento bibliográfico sobre os principais estudos e problemas relacionados ao polimorfismo em fármacos presentes nos medicamentos disponibilizados pela Farmácia Popular do Brasil. O polimorfismo deve ser controlado a fim de evitar possível ineficácia terapêutica e/ou dosagem inapropriada dos medicamentos. Destacamos que são poucos os ensaios obrigatórios para identificação e controle desse fenômeno em medicamentos, o que pode acarretar grande problema de saúde pública.