The work reports the impregnation of naproxen into locust bean gum mesoporous matrixes with different textural properties. The matrixes were prepared through the dissolution of the biopolymer in water and in two ionic liquids (ILs): [bmim][Cl] and [C2OHmim][Cl] and dried with scCO2. The poor water-soluble pharmaceutical drug naproxen was loaded into the matrixes and the composites were characterized by attenuated total reflectance-Fourier transform infrared spectroscopy and by differential scanning calorimetry; the results were compared with neat ILs and drug. The naproxen release from the matrixes was attempted at pH 7.4. Sustained release of naproxen in the different composites occurs, and consequently the naproxen release has lower rates compared with neat crystalline naproxen dissolution. Nevertheless, it was possible to observe small differences on release profiles for the studied composites. The higher release rate was observed for the composite where [bmim][Cl] was used as solvent, for which the calorimetric analysis revealed full amorphization of the incorporated drug. Cytotoxicity assays reveal that cellular viability in Caco-2 cells is preserved. This fact allied with the biocompatibility of locust bean gum allow for the composites potential application as naproxen controlled/sustained delivery systems with higher drug bioavailability achieved through naproxen amorphization.

Lyotropic colloidal aqueous suspensions of cellulose nanocrystals (CNCs) can, after solvent evaporation, retain their chiral nematic arrangement. As water is removed the pitch value of the suspension decreases and structural colour-generating films, which are mechanically brittle in nature, can be obtained. Increasing their flexibility while keeping the chiral nematic structure and biocompatible nature is a challenging task. However, if achievable, this will promote their use in new and interesting applications. In this study, we report on the addition of different amounts of hydroxypropyl cellulose (HPC) to CNCs suspension within the coexistence of the isotropic-anisotropic phases and infer the influence of this cellulosic derivative on the properties of the obtained solid films. It was possible to add 50 wt.% of HPC to a CNCs aqueous suspension (to obtain a 50/50 solids ratio) without disrupting the LC phase of CNCs and maintaining a left-handed helical structure in the obtained films. When 30 wt.% of HPC was added to the suspension of CNCs, a strong colouration in the film was still observed. This colour shifts to the near-infrared region as the HPC content in the colloidal suspension increases to 40 wt.% or 50 wt.% The all-cellulosic composite films present an increase in the maximum strain as the concentration of HPC increases, as shown by the bending experiments and an improvement in their thermal properties.

Abstract Anaerobic microorganisms of the Geobacter genus are effective electron sources for the synthesis of nanoparticles, for bioremediation of polluted water, and for the production of electricity in fuel cells. In multistep reactions, electrons are transferred via iron/heme cofactors of c-type cytochromes from the inner cell membrane to extracellular metal ions, which are bound to outer membrane cytochromes. We measured electron production and electron flux rates to 5×105 e s−1 per G. sulfurreducens. Remarkably, these rates are independent of the oxidants, and follow zero order kinetics. It turned out that the microorganisms regulate electron flux rates by increasing their Fe2+/Fe3+ ratios in the multiheme cytochromes whenever the activity of the extracellular metal oxidants is diminished. By this mechanism the respiration remains constant even when oxidizing conditions are changing. This homeostasis is a vital condition for living systems, and makes G. sulfurreducens a versatile electron source.

The monoheme outer membrane cytochrome F (OmcF) from Geobacter sulfurreducens plays an important role in Fe(III) reduction and electric current production. The electrochemical characterization of this cytochrome has shown that its redox potential is modulated by the solution pH (redox-Bohr effect) endowing the protein with the necessary properties to couple electron and proton transfer in the physiological range. The analysis of the OmcF structures in the reduced and oxidized states showed that with the exception of the side chain of histidine 47 (His⁴⁷), all other residues with protonatable side chains are distant from the heme iron and, therefore, are unlikely to affect the redox potential of the protein. The protonatable site at the imidazole ring of His⁴⁷ is in the close proximity to the heme and, therefore, this residue was suggested as the redox-Bohr center. In the present work, we tested this hypothesis by replacing the His⁴⁷ with non-protonatable residues (isoleucine – OmcFH47I and phenylalanine – OmcFH47F). The structure of the mutant OmcFH47I was determined by X-ray crystallography to 1.13 Å resolution and showed only minimal changes at the site of the mutation. Both mutants were ¹⁵N-labeled and their overall folding was confirmed to be the same as the wild-type by NMR spectroscopy. The pH dependence of the redox potential of the mutants was measured by cyclic voltammetry. Compared to the wild-type protein, the magnitude of the redox-Bohr effect in the mutants was smaller, but not fully abolished, confirming the role of His⁴⁷ on the pH modulation of OmcF’s redox potential. However, the pH effect on the heme substituents’ NMR chemical shifts suggested that the heme propionate P₁₃ also contributes to the overall redox-Bohr effect in OmcF. In physiological terms, the contribution of two independent acid–base centers to the observed redox-Bohr effect confers OmcF a higher versatility to environmental changes by coupling electron/proton transfer within a wider pH range.

Nanotechnology provides new tools for gene expression analysis that allow for sensitive and specific characterization of prognostic signatures related to cancer. Cancer is a complex disease where multiple gene loci contribute to the phenotype. The ability to simultaneously monitor differential expression originating from each locus allows for a more accurate indication into the degree of cancerous activity than either locus alone. Metal nanoparticles have been widely used as labels for in vitro identification and quantification of target sequences. Here we describe the synthesis of nanoparticles with different noble metal compositions in an alloy format that are then functionalized with thiol-modified ssDNA (nanoprobes). We also show how such nanoprobes are used in a non-cross-linking colorimetric method for the direct detection and quantification of specific mRNA targets, without the need for enzymatic amplification or reverse-transcription steps. The different metals in the alloy provide for distinct absorption spectra due to their characteristic plasmon resonance peaks. The color multiplexing allows for simultaneous identification of different mRNA targets involved in cancer development. A comparison of the absorption spectra of the nanoprobe mixtures taken before and after induced aggregation of metal nanoparticles allows to both identify and quantify each mRNA target. We describe the use of gold and gold–silver alloy nanoprobes for the development of the non-cross-linking method to detect a specific BCR-ABL fusion gene (e.g., e1a2 and e14a2) mRNA target associated with chronic myeloid leukemia (CML) using 10 ng/μL of unamplified total human RNA. Additionally, we demonstrate the use of this approach for the direct diagnostics of CML. This simple methodology takes less than 50 min to complete after total RNA extraction with comparable specificity and sensitivity to the more commonly used methods.

Biofilms are generally defined as communities of cells involved in a self-produced extracellular matrix adhered to a surface. In biofilms, the bacteria are less sensitive to host defense mechanisms and antimicrobial agents, due to multiple strategies, that involve modulation of gene expression, controlled metabolic rate, intercellular communication, composition, and 3D architecture of the extracellular matrix. These factors play a key role in streptococci pathogenesis, contributing to therapy failure and promoting persistent infections. The species of the pyogenic group together with Streptococcus pneumoniae are the major pathogens belonging the genus Streptococcus, and its biofilm growth has been investigated, but insights in the genetic origin of biofilm formation are limited. This review summarizes pyogenic streptococci biofilms with details on constitution, formation, and virulence factors associated with formation.

The bacterium Geobacter metallireducens is highly efficient in long-range extracellular electron transfer, a process that relies on an efficient bridging between the cytoplasmic electron donors and the extracellular acceptors. The periplasmic triheme cytochromes are crucial players in these processes and thus the understanding of their functional mechanism is crucial to elucidate the extracellular electron transfer processes in this microorganism. The triheme cytochrome PpcF from G. metallireducens has the lowest amino acid sequence identity with the remaining cytochromes from the PpcA-family of G. sulfurreducens and G. metallireducens, making it an interesting target for structural and functional studies. In this work, we performed a detailed functional and thermodynamic characterization of cytochrome PpcF by the complementary usage of NMR and visible spectroscopic techniques. The results obtained show that the heme reduction potentials are negative, different from each other and are also modulated by the redox and redox-Bohr interactions that assure unprecedented mechanistic features to the protein. The results showed that the order of oxidation of the hemes in cytochrome PpcF is maintained in the entire physiological pH range. The considerable separation of the hemes' redox potential values facilitates a sequential transfer within the chain of redox centers in PpcF, thus assuring electron transfer directionality to the electron acceptors.

Among the movements observed in some cellulosic structures produced by plants are those that involve the dispersion and burial of seeds, as for example in Erodium from the Geraniaceae plant family. Here we report on a simple and efficient strategy to isolate and tune cellulose-based hygroscopic responsive materials from Erodium awns' dead tissues. The stimuli-responsive material isolated forms left-handed (L) or right-handed (R) helical birefringent transparent ribbons in the wet state that reversibly change to R helices when the material dries. The humidity-driven motion of dead tissues is most likely due to a composite material made of cellulose networks of fibrils imprinted by the plant at the nanoscale, which reinforces a soft wall polysaccharide matrix. The inversion of the handedness is explained using computational simulations considering filaments that contract and expand asymmetrically. The awns of Erodium are known to present hygroscopic movements, forming R helices in the dry state, but the possibility of actuating chirality via humidity suggests that these cellulose-based skeletons, which do not require complicated lithography and intricate deposition techniques, provide a diverse range of applications from intelligent textiles to micro-machines.

The syntheses of the heterometallic sodium and potassium-dioxidovanadium 2D polymers, [NaVO2(1κNOO’;2κO”-L)(H2O)]n (1) and [KVO2(1κNOO’;2κO’;3κO”-L)(EtOH)]n (2) (where the κ notation indicates the coordinating atoms of the polydentate ligand L) derived from (3,5-di-tert-butyl-2-hydroxybenzylidene)-2-hydroxybenzohydrazide (H2L) are reported. The polymers were characterized by IR, NMR, elemental analysis and single crystal X-ray diffraction analysis. The antiproliferative potential of 1 and 2 was examined towards four human cancer cell lines (ovarian carcinoma, A2780, colorectal carcinoma, HCT116, prostate carcinoma, PC3 and breast adenocarcinoma, MCF-7cell lines) and normal human fibroblasts. Complex 1 and 2 showed the highest cytotoxic activity against A2780 cell line (IC50 8.2 and 11.3 μM, respectively) with 1 > 2 and an IC50 in the same range as cisplatin (IC50 3.4 μM; obtained in the same experimental conditions) but, interestingly, with no cytotoxicity to healthy human fibroblasts for concentrations up to 75 μM. This high cytotoxicity of 1 in ovarian cancer cells and its low cytotoxicity in healthy cells demonstrates its potential for further biological studies. Our results suggest that both complexes induce ovarian carcinoma cell death via apoptosis and autophagy, but autophagy is the main biological cause of the reduction of viability observed and that ROS (reactive oxygen species) may play an important role in triggering cell death.

The materials described in this work result from the selfassembly of liquid crystals and ionic liquids into droplets,
stabilized within a biopolymeric matrix. These systems are
extremely versatile gels, in terms of composition, and offer
potential for fine tuning of both structure and function, as
each individual component can be varied. Here, the
characterization and application of these gels as sensing thin
films in gas sensor devices is presented. The unique
supramolecular structure of the gels is explored for molecular
recognition of volatile organic compounds (VOCs) by
employing gels with distinct formulations to yield
combinatorial optical and electrical responses used in the
distinction and identification of VOCs.

Herein we report the synthesis of novel ionic liquids (ILs) and organic salts by combining ibuprofen as anion with ammonium, imidazolium, or pyridinium cations. The methodology consists of an acid–base reaction of neutral ibuprofen with cation hydroxides, which were previously prepared by anion exchange from the corresponding halide salts with Amberlyst A-26(OH). In comparison with the parent drug, these organic salts display higher solubility in water and biological fluids and a smaller degree of polymorphism, which in some cases was completely eliminated. With the exception of [C 16 Pyr][Ibu] and [N 1,1,2,2OH1 ][Ibu], the prepared salts did not affect the viability of normal human dermal fibroblasts or ovarian carcinoma (A2780) cells. Therefore, these ibuprofen-based ionic liquids may be very promising lead candidates for the development of effective formulations of this drug.

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While screening for polyhydroxyalkanoate (PHA) producing strains, using glycerol rich by-product as carbon source, it was observed that extracellular polymers were also secreted into the culture broth. The scope of this study was to characterize both intracellular and extracellular polymers, produced by Pseudomonas putida NRRL B-14875 and Pseudomonas chlororaphis DSM 50083, mostly focusing on those novel extracellular polymers. It was found that they fall into the class of bioemulsifiers (BE), as they showed excellent emulsion stability against different hydrocarbons/oils at various pH conditions, temperature and salinity concentrations. Cytotoxicity tests revealed that BE produced by P. chlororaphis inhibited the growth of highly pigmented human melanoma cells (MNT-1) by 50% at concentrations between 150 and 200 μg/mL, while no effect was observed on normal skin primary keratinocytes and melanocytes. This is the first study reporting mcl-PHA production by P. putida NRRL B-14785 and bioemulsifier production from both P. putida and P. chlororaphis strains.

Anthocyanins may protect against a myriad of human diseases. However few studies have been conducted to evaluate their bioavailability so their absorption mechanism remains unclear. This study aimed to evaluate the role of two glucose transporters (GLUT1 and GLUT3) in anthocyanins absorption in the human gastric epithelial cells (MKN-28) by using gold nanoparticles to silence these transporters. Anthocyanins were purified from purple fleshed sweet potatoes and grape skin. Silencing of GLUT1 and/or GLUT3 mRNA was performed by adding AuNP@GLUT1 and/or AuNP@GLUT3 to MKN-28 cells. Downregulation of mRNA expression occurred concomitantly with the reduction in protein expression. Malvidin-3-O-glucoside (Mv3glc) transport was reduced in the presence of either AuNP@GLUT1 and AuNP@GLUT3, and when both transporters were blocked simultaneously. Peonidin-3-(6′-hydroxybenzoyl)-sophoroside-5-glucoside (Pn3HBsoph5glc) and Peonidin-3-(6′-hydroxybenzoyl-6″-caffeoyl)-sophoroside-5-glucoside (Pn3HBCsoph5glc) were assayed to verify the effect of the sugar moiety esterification at glucose B in transporter binding. Both pigments were transported with a lower transport efficiency compared to Mv3glc, probably due to steric hindrance of the more complex structures. Interestingly, for Pn3HBCsoph5glc although the only free glucose is at C5 and the inhibitory effect of the nanoparticles was also observed, reinforcing the importance of glucose on the transport regardless of its position or substitution pattern. The results support the involvement of GLUT1 and GLUT3 in the gastric absorption of anthocyanins.

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The major challenge of the pharmaceutical industry is to find potential solvents for poorly water-soluble drug molecules. Ionic liquids (ILs) have attracted this industry as (co-) solvents due to their unique physicochemical and biological properties. Herein, a straightforward approach for the enhancement of water solubility of paracetamol and sodium diclofenac is presented, using new biocompatible N-acetyl amino acid N-alkyl cholinium-based ionic liquids as co-solvents (0.2 - 1 mol%). These new ionic liquids were able to increase water solubility of these drugs up to four times higher than in pure water or in an inorganic salt solution. In the presence of these ILs the drugs lipophilicity (log Kow) was not significantly changed for paracetamol, but for sodium diclofenac it was possible to decrease significantly its lipophilicity. Concerning cytotoxicity in human dermal fibroblasts it was observed that ILs did not show a significant toxicity, and were able to improve cell viability compared with the respective precursors.

The syntheses of the heterometallic sodium and potassium-dioxidovanadium 2D polymers, [NaVO2(1kappaNOO';2kappaO"-L)(H2O)]n(1) and [KVO2(1kappaNOO';2kappaO';3kappaO"-L)(EtOH)]n(2) (where the kappa notation indicates the coordinating atoms of the polydentate ligand L) derived from (3,5-di-tert-butyl-2-hydroxybenzylidene)-2-hydroxybenzohydrazide (H2L) are reported. The polymers were characterized by IR, NMR, elemental analysis and single crystal X-ray diffraction analysis. The antiproliferative potential of 1 and 2 was examined towards four human cancer cell lines (ovarian carcinoma, A2780, colorectal carcinoma, HCT116, prostate carcinoma, PC3 and breast adenocarcinoma, MCF-7cell lines) and normal human fibroblasts. Complex 1 and 2 showed the highest cytotoxic activity against A2780 cell line (IC50 8.2 and 11.3muM, respectively) with 1>2 and an IC50 in the same range as cisplatin (IC50 3.4muM; obtained in the same experimental conditions) but, interestingly, with no cytotoxicity to healthy human fibroblasts for concentrations up to 75muM. This high cytotoxicity of 1 in ovarian cancer cells and its low cytotoxicity in healthy cells demonstrates its potential for further biological studies. Our results suggest that both complexes induce ovarian carcinoma cell death via apoptosis and autophagy, but autophagy is the main biological cause of the reduction of viability observed and that ROS (reactive oxygen species) may play an important role in triggering cell death.

Microbial electrosynthesis is an emerging green technology that explores the capability of a particular group of microorganisms to drive their metabolism toward the production of hydrogen or value-added chemicals from electrons supplied by electrode surfaces. The cytochrome PccH showed the largest increase in transcription when electrons are supplied to Geobacter sulfurreducens biofilms. Gene knock-out experiments have shown that the electron transfer toward G. sulfurreducens cells was completely inhibited by the deletion of the gene encoding for cytochrome PccH. This identifies a crucial role for this protein in G. sulfurreducens microbial electrosynthesis mechanisms, which are currently unknown. In this work, we present the backbone (1H, 13C and 15N) and heme assignment for PccH in the oxidized state. The data obtained paves the way to identify and structurally map the molecular interaction regions between the cytochrome PccH and its physiological redox partners.

Streptococcus dysgalactiae subsp. dysgalactiae (SDSD), a Lancefield group C streptococci (GCS), is a frequent cause of bovine mastitis. This highly prevalent disease is the costliest in dairy industry. Adherence and biofilm production are important factors in streptoccocal pathogenesis. We have previously described the adhesion and internalization of SDSD isolates in human cells and now we describe the biofilm production capability of this bacterium. In this work we integrated microbiology, imaging and computational methods to evaluate the biofilm production capability of SDSD isolates; to assess the presence of biofilm regulatory protein BrpA homolog in the biofilm producers; and to predict a structural model of BrpA-like protein and its binding to putative inhibitors. Our results show that SDSD isolates form biofilms on abiotic surface such as glass (hydrophilic) and polystyrene (hydrophobic), with the strongest biofilm formation observed in glass. This ability was mainly associated with a proteinaceous extracellular matrix, confirmed by the dispersion of the biofilms after proteinase K and trypsin treatment. The biofilm formation in SDSD isolates was also confirmed by confocal laser scanning microscopy (CLSM) and scanning electron microscopy (SEM). Under SEM observation, VSD16 isolate formed cell aggregates during biofilm growth while VSD9 and VSD10 formed smooth and filmy layers. We show that brpA-like gene is present and expressed in SDSD biofilm-producing isolates and its expression levels correlated with the biofilm production capability, being more expressed in the late exponential phase of planktonic growth compared to biofilm growth. Fisetin, a known biofilm inhibitor and a putative BrpA binding molecule, dramatically inhibited biofilm formation by the SDSD isolates but did not affect planktonic growth, at the tested concentrations. Homology modeling was used to predict the 3D structure of BrpA-like protein. Using high throughput virtual screening and molecular docking, we selected five ligand molecules with strong binding affinity to the hydrophobic cleft of the protein, making them potential inhibitor candidates of the SDSD BrpA-like protein. These results warrant further investigations for developing novel strategies for SDSD anti-biofilm therapy.

A series of 2,2':6',2''-terpyridine (terpy), 2,6-di(thiazol-2-yl)pyridine (dtpy) and 2,6-di(pyrazin-2-yl)pyridine (dppy) derivatives with n-quinolyl substituents (n = 2 and 4) was used to synthesize five-coordinate complexes [CuCl2(n-quinolyl-terpy)] (1-2), [CuCl2(n-quinolyl-dtpy)] (3-4) and [CuCl2(n-quinolyl-dppy)] (5-6), respectively. The main emphasis of the research was to investigate the impact of the triimine skeleton (terpy, dtpy and dppy) and n-quinolyl pendant substituent on the antiproliferative and catalytic properties of 1-6. The obtained Cu(ii) compounds were studied as antiproliferative agents against human colorectal (HCT116) and ovarian (A2780) carcinoma, and they were used as catalysts for the oxidation of alkanes and alcohols with peroxides under mild conditions. The kinetic characteristics of the oxidizing species generated by the catalytic system Cu(ii) complex-H2O2 in CH3CN were obtained from the dependence of the alkane oxidation rate on its initial concentration. A model of competitive interaction of hydroxyl radicals with CH3CN and RH in the catalyst cavity has been proposed which is based on the simultaneous study of kinetics and selectivity in alkane oxidations.

The major challenge of the pharmaceutical industry is to find potential solvents for poorly water-soluble drug molecules. Ionic liquids (ILs) have attracted this industry as (co-) solvents due to their unique physicochemical and biological properties. Herein, a straightforward approach for the enhancement of the water solubility of paracetamol and sodium diclofenac is presented, using new biocompatible N-acetyl amino acid N-alkyl cholinium-based ionic liquids as co-solvents (0.2-1mol%). These new ionic liquids were able to increase the water solubility of these drugs up to four times that in pure water or in an inorganic salt solution. In the presence of these ILs, the drugs lipophilicity (log P was not significantly changed for paracetamol, but for sodium diclofenac it was possible to decrease significantly its lipophilicity. Concerning cytotoxicity in human dermal fibroblasts it was observed that ILs did not show a significant toxicity, and were able to improve cell viability compared with the respective precursors.