Uncomplexed Form Research Articles

Pulmonary delivery of forsythin-phospholipid complexes improves the lung anti-inflammatory efficacy in mice by enhancing dissolution and lung tissue affinity

Forsythin, currently in phase II clinical trials in China for the treatment of the common cold and influenza, faces challenges in achieving adequate lung drug exposure due to its limited dissolution and permeability, thereby restricting its therapeutic efficacy. The objective of this work was to formulate a forsythin-phospholipid complex (FPC) to enhance its dissolution properties and lung affinity with a particular view to improving pulmonary drug exposure and anti-inflammatory response. The results revealed that forsythin reacted with dipalmitoyl-phosphatidylcholine to form a stable, nanosized FPC suspension. This formulation significantly improved the in vitro drug's dissolution, cellular uptake, and lung affinity compared to its uncomplexed form. Intratracheal administration of FPC in a mouse model of acute lung injury induced by lipopolysaccharide (LPS) resulted in a substantial increase in drug exposure to lung tissues (39.6-fold) and immune cells in the epithelial lining fluid (198-fold) compared to intraperitoneal injection. In addition, FPC instillation exhibited superior local anti-inflammatory effects, leading to improved survival rates among mice with LPS-induced acute respiratory distress syndrome, outperforming both instilled forsythin and injected FPC. Overall, this work demonstrated the potential of phospholipid complexes as a viable option for developing inhalation products for drugs with limited solubility and permeability properties.

Calix[4]pyrrolato-germane-(thf)2: Unlocking the Anti-van't Hoff-Le Bel Reactivity of Germanium(IV) by Ligand Dissociation.

Anti-van't Hoff-Le Bel configured p-block element species possess intrinsically high reactivity and are thus challenging to isolate. Consequently, numerous elements in this configuration, including square-planar germanium(IV), remain unexplored. Herein, we follow a concept to reach anti-van't Hoff-Le Bel reactivity by ligand dissociation from a rigid calix[4]pyrrole germane in its bis(thf) adduct. While the macrocyclic ligand assures square-planar coordination in the uncomplexed form, the labile thf donors provide robustness for isolation on a multigram scale. Unique properties of a low-lying acceptor orbital imparted to germanium(IV) can be verified, e.g., by isolating an elusive anionic hydrido germanate and exploiting it for challenging bond activations. Aldehydes, water, alcohol, and a CN triple bond are activated for the first time by germanium-ligand cooperativity. Unexpected behaviors against fluoride ion donors disclose critical interferences of a putative redox-coupled fluoride ion transfer during the experimental determination of Lewis acidity. Overall, we showcase how ligand lability grants access to the uncharted chemistry of anti-van't Hoff-Le Bel germanium(IV) and line up this element as a member in the emerging class of structurally constrained p-block elements.

Stability screening of pharmaceutical cocrystals

An efficient protocol for assessing both the chemical and physical stability of cocrystalline forms of active pharmaceutical ingredients (APIs) is proposed. In this protocol, the cocrystalline material is used to prepare two standard formulations, mimicking wet granulations, to make low-dose tablets. After designed stress testing at a range of temperatures and RH conditions, degradant formation is modeled from the data using ASAPprime ® to determine if the tablets have a minimum of a one-year shelf-life (25 °C/60% RH open). When the cocrystals provide a kinetic solubility enhancement over the un-complexed API, a physical assessment of the cocrystal stability is carried out using the same tablets at selected stress conditions. For this assessment, kinetic solubility (where the amount of buffer used to dissolve the tablet is adjusted to completely dissolve the cocrystalline form but leave most of the un-complexed form out of solution) changes are used to indicate whether there is a significant risk for physical instability on long-term storage. This process was exemplified using model cocrystals of APIs.

Antihypertensive effect of carvacrol is improved after incorporation in β-cyclodextrin as a drug delivery system.

Carvacrol (CARV), has been shown to possess various pharmacological properties, especially in the treatment of cardiovascular diseases. We evaluated the antihypertensive effect of the CARV free and encapsulation of CARV in β-cyclodextrin (CARV/β-CD), and whether CARV/β-CD is able to improve the antihypertensive effects of CARV free in spontaneously hypertensive rats (SHR). The rats were randomly divided into four groups, each treated daily for 21days and the mean arterial pressure and heart rate was measured every 5days: group 1, Wistar-vehicle solution; group 2, SHR-vehicle; group 3, SHR-CARV 50mg/kg/d; and group 4, CARV/β-CD 50mg/kg/d. After 21days of treatment, the mesenteric artery from treated animals was tested for phenylephrine (Phe) and sodium nitroprusside (SNP) sensitivity. In addition, administration of CARV/β-CD induced important antihypertensive activity when compared with the uncomplexed form, reducing the progression of arterial hypertension in SHR. Moreover, pharmacological potency to Phe in the SHR-CARV and CARV/β-CD groups was increased, approaching values expressed in the Wistar-vehicle. Furthermore, CARV/β-CD reduced the production of the pro-inflammatory mediator, IL-1β, and increased anti-inflammatory cytokine, IL-10. Together, these results produced evidence that the encapsulation of CARV in β-CD can improve cardiovascular activity, showing potential anti-inflammatory and antihypertensive effects.

Structural analysis of saccharin in aqueous solution by NMR and supramolecular interactions with α-, β-, γ-cyclodextrins

Abstract As one of the many sweet-tasting molecules of various sizes and chemical/physical properties, interaction of saccharin with water molecules, and α-, β-, and γ-cyclodextrin in aqueous solution, is examined using NMR properties of both exchangeable and non-exchangeable protons. A thorough assignment of 13C and 1H signals of saccharin is achieved and corresponding precise chemical shifts were reported. Direct complexation interaction with α-cyclodextrin is absent, whereas partial interaction with β-cyclodextrin and full immersion/complexation with γ-cyclodextrin take place with reference to the NMR parameters including complexation induced chemical shift (CIS) changes, temperature coefficients, and exchange rates. The interaction with β-cyclodextrin seems to be taking place on the surface without full insertion into cavity, resulting in only slight chemical shift changes of maximum |0.017| ppm on saccharin proton signals with the signal patterns almost identically similar to uncomplexed form. This superficial interaction does, nevertheless, have effect on CIS values of β-cyclodextrin hydroxy protons with a maximum of |0.091| ppm. For γ-cyclodextrin, the full complexation interaction is directly visible from the large CIS of C(5)H and C(8)H protons of saccharin, as well as those of O(2)H and O(6)H of γ-cyclodextrin.

Modulation of photochromic properties of spirooxazine bearing sulfobutyl substituent by metal ions

A novel spirooxazine bearing sulfobutyl substituent on the indoline part of the molecule has been synthesized and its photochromic properties and complex formation with metal ions have been studied. Under UV light irradiation of colorless acetonitrile solution of spirooxazine the open merocyanine form having an absorption band in visible region of the spectrum with λmax = 625 nm is generated. Thermal back reaction of merocyanine form to spirooxazine with the rate constant 1.10 s−1 at 298 K has been observed. Interaction of spirooxazine with Mg2+, Ca2+, Cd2+, Zn2+ or Pb2+ ions in the dark leads to the formation of complexes of merocyanine form demonstrating a visible absorption band shifted from 20 to 8 nm relative to the uncomplexed merocyanine form. The activation energy of metal-induced coloration process in the dark is in the range of 84–88 kJ mol−1 and does not depend on the nature of the metal ions. This suggests that the process of thermal ring opening of the spiro form is the rate limiting step of complex formation. The degree of conversion of spirooxazine to complexed merocyanine form strongly depends on metal ions nature and irradiation wavelength.

Antihypertensive potential of linalool and linalool complexed with β-cyclodextrin: Effects of subchronic treatment on blood pressure and vascular reactivity

Linalool (LIN) is a monoterpene alcohol present in some aromatic medicinal plants with biological activities that can impact cardiovascular diseases. This chemical class is highly volatile and β-cyclodextrin (β-CD) has been employed to improve the pharmacological properties of monoterpenes. Thus, the aim of this study was to evaluate the cardiovascular effects of LIN free focusing on the antihypertensive properties of this monoterpene and to study whether LIN, complexed in β-cyclodextrin (LIN-βCD) is able to improve the pharmacological activity of LIN. Spontaneously hypertensive rats (SHR) were randomly divided into 5 groups, each treated daily for 21 days, in the following manner: group 1 (vehicle solution); group 2 (captopril; 30 mg/kg/day); group 3 (LIN; 100 mg/kg/day); group 4 (LIN; 50 mg/kg/day) and group 5 (LIN/β-CD; 50 mg/kg/day). Daily body weight measurements were conducted and mean arterial pressure and heart rate were measured every 5 days. The mesenteric artery from treated animals was tested for phenylephrine and sodium nitroprusside (SNP) sensitivity. The SHR treated with vehicle demonstrated progressive increase in mean arterial pressure and captopril, a positive control, induced a significant decrease. After 21 days of treatment, the blood pressure of the SHR treated by (−)-LIN (100 mg/kg) was significantly reduced. In addition, various important cardiovascular parameters improved, including: the treatment with LIN prevented the development of cardiac hypertrophy, increased levels of the anti-inflammatory cytokine (IL-10), increased vasodilator responsiveness and reduced sensitivity to the sympathetic agonist. Furthermore, the inclusion complex containing LIN in β-CD produced a higher antihypertensive profile when compared with uncomplexed form. Taking together, our results suggested that LIN shown a potential antihypertensive effect and β-CD may be an important tool to improve the cardiovascular activity of LIN and other water-insoluble compounds.

D-limonene exhibits superior antihyperalgesic effects in a β-cyclodextrin-complexed form in chronic musculoskeletal pain reducing Fos protein expression on spinal cord in mice

Chronic musculoskeletal pain is one of the main symptoms found in Fibromyalgia with unclear etiology and limited pharmacological treatment. The aim of this study was to complex LIM in β-cyclodextrin (LIM-βCD) and then evaluate its antihyperalgesic effect in an animal model of chronic musculoskeletal pain. Differential scanning calorimetry and scanning electron microscopy was used for the characterization of the inclusion complex. Male Swiss mice were used for experimental procedures where mechanical hyperalgesia, thermal hyperalgesia, muscular strength, Fos immunofluorescence was studied after induction of hyperalgesia. Mechanism of action was also investigated through tail flick test and capsaicin-induced nociception. Endothermic events and morphological changes showed that the slurry complex method was the best method for the complexation. After induction of hyperalgesia, the oral administration of LIM-βCD (50mg/kg) significantly increased the paw withdrawal threshold compared to uncomplexed limonene. Fos immunofluorescence showed that both compounds significantly decreased the number of Fos-positive cells in the dorsal horn. In nociceptive tests, FLU was able to reverse the antinociceptive effect of LIM-βCD. After intraplantar administration of capsaicin, LIM was able to significantly decrease time to lick. LIM-βCD has antihyperalgesic action superior to its uncomplexed form, with possible action in the dorsal horn of the spinal cord. These results suggest the possible applicability of LIM, uncomplexed or complexed with βCD, in conditions such as FM and neuropathic pain, for which there are currently only limited pharmacological options.

Heteroatom Tuning of Bimolecular Criegee Reactions and Its Implications

AbstractHigh‐level quantum‐chemical calculations have been performed to understand the key reactivity determinants of bimolecular reactions of Criegee intermediates and H2X (X=O, S, Se, and Te). Criegee intermediates are implicated as key intermediates in atmospheric, synthetic organic, and enzymatic chemistry. Generally, it is believed that the nature and location of substituents at the carbon of the Criegee intermediate play a key role in determing the reactivity. However, the present work suggests that it is not only the substitution of the Criegee intermediate, but the nature of the heteroatom in H2X that also plays a crucial role in determining the reactivity of the interaction between the Criegee intermediate and H2X. The barriers for the reactions of Criegee intermediates and H2X satisfy an inverse correlation with the bond strength of X−H in H2X, and a direct correlation with the first pKa of H2X. This heteroatom tuning causes a substantial barrier lowering of 8–11 kcal mol−1 in the Criegee reaction barrier in going from H2O to H2Te. An important implication of these results is that the reaction of the Criegee intermediate and H2S could be a source of thioaldehydes, which are important in plantery atmospheres and synthetic organic chemistry. By performing the reaction of Criegee intermediates and H2S under water or acid catalysis, thioladehydes could be detected in a hydrogen‐bonded complexed state, which is significantly more stable than their uncomplexed form. As a result, simpler aliphatic thioaldehydes could be selectively synthesized in the laboratory, which, otherwise, has been a significant synthetic challenge because of their ability to oligomerize.

Heteroatom Tuning of Bimolecular Criegee Reactions and Its Implications.

High-level quantum-chemical calculations have been performed to understand the key reactivity determinants of bimolecular reactions of Criegee intermediates and H2 X (X=O, S, Se, and Te). Criegee intermediates are implicated as key intermediates in atmospheric, synthetic organic, and enzymatic chemistry. Generally, it is believed that the nature and location of substituents at the carbon of the Criegee intermediate play a key role in determing the reactivity. However, the present work suggests that it is not only the substitution of the Criegee intermediate, but the nature of the heteroatom in H2 X that also plays a crucial role in determining the reactivity of the interaction between the Criegee intermediate and H2 X. The barriers for the reactions of Criegee intermediates and H2 X satisfy an inverse correlation with the bond strength of X-H in H2 X, and a direct correlation with the first pKa of H2 X. This heteroatom tuning causes a substantial barrier lowering of 8-11 kcal mol-1 in the Criegee reaction barrier in going from H2 O to H2 Te. An important implication of these results is that the reaction of the Criegee intermediate and H2 S could be a source of thioaldehydes, which are important in plantery atmospheres and synthetic organic chemistry. By performing the reaction of Criegee intermediates and H2 S under water or acid catalysis, thioladehydes could be detected in a hydrogen-bonded complexed state, which is significantly more stable than their uncomplexed form. As a result, simpler aliphatic thioaldehydes could be selectively synthesized in the laboratory, which, otherwise, has been a significant synthetic challenge because of their ability to oligomerize.

Modeling and fitting protein-protein complexes to predict change of binding energy.

It is possible to accurately and economically predict change in protein-protein interaction energy upon mutation (ΔΔG), when a high-resolution structure of the complex is available. This is of growing usefulness for design of high-affinity or otherwise modified binding proteins for therapeutic, diagnostic, industrial, and basic science applications. Recently the field has begun to pursue ΔΔG prediction for homology modeled complexes, but so far this has worked mostly for cases of high sequence identity. If the interacting proteins have been crystallized in free (uncomplexed) form, in a majority of cases it is possible to find a structurally similar complex which can be used as the basis for template-based modeling. We describe how to use MMB to create such models, and then use them to predict ΔΔG, using a dataset consisting of free target structures, co-crystallized template complexes with sequence identify with respect to the targets as low as 44%, and experimental ΔΔG measurements. We obtain similar results by fitting to a low-resolution Cryo-EM density map. Results suggest that other structural constraints may lead to a similar outcome, making the method even more broadly applicable.

Abstract P1-16-08: Inhibition of β-catenin pathway to overcome endocrine resistance in breast cancer

Abstract Background: It is well known that Wnt/β-catenin signaling pathway is involved in hormone receptor negative breast cancer, especially triple-negative breast cancer. In the case of hormone resistant breast cancer cells, they rarely showed hormone receptor expression. So we aimed to investigate whether the β-catenin pathway becomes activated in endocrine-resistant breast cancer cells and inhibition of β-catenin pathway can overcome endocrine resistance. Methods: We have established an MCF-7-derived tamoxifen-resistant cell line (TamR) by long-term culture of MCF-7 cells with gradually increasing 4-hydroxytamoxifen concentration till 3μM. The levels of protein expression and mRNA transcripts were determined using western blot analysis and real-time quantitative PCR. The transcriptional activity of β-catenin was measured using luciferase activity assay. We used ICG-001 as inhibitor of β-catenin transcription activity. Results: The expression of estrogen receptor was significantly decreased in TamR cells. On the other hand, the expressions of HER2 and EGFR were increased in TamR cells than in control cells. The active (uncomplexed form) β-catenin level was increased in TamR cells, and also showed a significantly increased β-catenin transcriptional activity. The ICG-001, small-molecular inhibitor of Wnt/β-catenin pathway, treatment significantly reduced β-catenin transcriptional activity in TamR cells. The ICG-001 reduced cell viability fo TamR cells which showed resistance to tamoxifen by 65.3%, and also inhibited target gene cyclin D1 expression. The combination of ICG-001 and mTOR inhibitor, rapamycin reduced cell viability of TamR cells by 81.7% and there was an additive effect of two drugs as a combination index of 1.022. Conclusions: The β-catenin pathway is activated in endocrine-resistant breast cancer cells and inhibition of that pathway would be a new therapeutic strategy which overcomes endocrine resistance in breast cancer. Citation Format: Won HS, Lee KE, Lee KM, Nam EM, Mun Y-C, Seong C-M, Lee SN. Inhibition of β-catenin pathway to overcome endocrine resistance in breast cancer. [abstract]. In: Proceedings of the Thirty-Eighth Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2015 Dec 8-12; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2016;76(4 Suppl):Abstract nr P1-16-08.

Characteristics of Sucrose Transport through the Sucrose-Specific Porin ScrY Studied by Molecular Dynamics Simulations.

Sucrose-specific porin (ScrY) is a transmembrane protein that allows for the uptake of sucrose under growth-limiting conditions. The crystal structure of ScrY was resolved before by X-ray crystallography, both in its uncomplexed form and with bound sucrose. However, little is known about the molecular characteristics of the transport mechanism of ScrY. To date, there has not yet been any clear demonstration for sucrose transport through the ScrY. Here, the dynamics of the ScrY trimer embedded in a phospholipid bilayer as well as the characteristics of sucrose translocation were investigated by means of atomistic molecular dynamics (MD) simulations. The potential of mean force (PMF) for sucrose translocation through the pore showed two main energy barriers within the constriction region of ScrY. Energy decomposition allowed to pinpoint three aspartic acids as key residues opposing the passage of sucrose, all located within the L3 loop. Mutation of two aspartic acids to uncharged residues resulted in an accordingly modified electrostatics and decreased PMF barrier. The chosen methodology and results will aid in the design of porins with modified transport specificities.

Abstract B49: Inhibition of β-catenin pathway to overcome endocrine resistance in tamoxifen-resistant breast cancer cell line

Abstract Background: It is well known that Wnt/β-catenin signaling pathway is involved in hormone receptor negative breast cancer, especially triple-negative breast cancer. In the case of hormone resistant breast cancer cells, they rarely showed hormone receptor expression. So we aimed to investigate whether the β-catenin pathway becomes activated in endocrine-resistant breast cancer cells and inhibition of β-catenin pathway can overcome endocrine resistance. Methods: We have established an MCF-7-derived tamoxifen-resistant cell line (TamR) by long-term culture of MCF-7 cells with gradually increasing 4-hydroxytamoxifen concentration till 3μM. The levels of protein expression and mRNA transcripts were determined using western blot analysis and real-time quantitative PCR. The transcriptional activity of β-catenin was measured using luciferase activity assay. We used ICG-001 as inhibitor of β-catenin transcription activity. Results: The expression of estrogen receptor was significantly decreased in TamR cells. On the other hand, the expressions of HER2 and EGFR were increased in TamR cells than in control cells. The active (uncomplexed form) β-catenin level was increased in TamR cells, and also showed a significantly increased β-catenin transcriptional activity. The ICG-001, small-molecular inhibitor of Wnt/β-catenin pathway, treatment significantly reduced β-catenin transcriptional activity in TamR cells. The ICG-001 reduced cell viability fo TamR cells which showed resistance to tamoxifen by 65.3%, and also inhibited target gene cyclin D1 expression. The combination of ICG-001 and mTOR inhibitor, rapamycin reduced cell viability of TamR cells by 81.7% and there was an additive effect of two drugs as a combination index of 1.022. Conclusions: The β-catenin pathway is activated in endocrine-resistant breast cancer cells and inhibition of that pathway would be a new therapeutic strategy which overcomes endocrine resistance in breast cancer. Citation Format: Hye Sung Won, Kyoung Eun Lee, Eun Mi Nam, Yeung-Chul Mun, Chu-Myong Seong, Soon Nam Lee. Inhibition of β-catenin pathway to overcome endocrine resistance in tamoxifen-resistant breast cancer cell line. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr B49.

Studies on UV–vis and fluorescence changements in Co2+ and Cu2+ recognition by a new benzimidazole–benzothiadiazole derivative

A novel fluorescent sensor (BI-T) was synthesized by the coupling reaction of 2-(2-hydroxyphenyl)-7-phenyl-1H-benzimidazole-4-boronic acid with 4,7-dibromo-2,1,3-benzothiadiazole via Pd-catalysed Suzuki reaction. Its photophysical properties were investigated systematically in different solvents to analyze its potential usage in the fluorescence detection for transition metal ions such as Co2+ and Cu2+ in solutions. In benzonitrile solution, new absorption bands for complex structure of BI-T at 470nm and 580nm were appeared in the presence of Co2+ ions with respect to the uncomplexed form of the sensor absorption. BI-T sensor is usable for both Co2+ and Cu2+ sensation through “on–off” fluorescence change in benzonitrile. In ethanol solution, BI-T shows similar optical performances to detect Co2+ and Cu2+ ions. BI-T detects trace amount of transition metals in ethanol with estimated limit of detection around 4.1×10−7M and 5.5×10−7M for Co2+ and Cu2+ ions, respectively. Possible formation of metal-to-ligand charge transfer (MLCT) state during the titration with Co2+ ion is supported by the detection of long-lived species in the excited state.

MicroRNAs as biomarkers in pituitary tumors.

The use of extracellular microRNAs (miRNAs) as circulating biomarkers is currently leading to relevant advances in the diagnosis and assessment of prognosis of several diseases. Specific miRNAs have also been shown to play a role in the pathophysiology of many neoplastic and non-neoplastic diseases. A number of studies have demonstrated that miRNAs show differential expression in various tumors, such as in the prostate, ovary, lung, breast, brain, and pituitary. Recent findings have built connections between miRNAs that are deregulated within the tumor and their presence in peripheral blood. MiRNAs have been shown to be stable in the blood where they are present in either free and/or uncomplexed form, as well as packed in microvesicles, exosomes, and apoptotic bodies, or bound to different proteins. Because the pituitary is a highly vascularized organ that releases hormones into the circulation, miRNAs would be useful biomarkers for the diagnosis of pituitary tumors, as well as for predicting or detecting recurrence after surgery. Here we review the biological significance of miRNAs in pituitary tumors and the potential value of circulating miRNAs as biomarkers.

Structural Context of Disease-Associated Mutations and Putative Mechanism of Autoinhibition Revealed by X-Ray Crystallographic Analysis of the EZH2-SET Domain

The enhancer-of-zeste homolog 2 (EZH2) gene product is an 87 kDa polycomb group (PcG) protein containing a C-terminal methyltransferase SET domain. EZH2, along with binding partners, i.e., EED and SUZ12, upon which it is dependent for activity forms the core of the polycomb repressive complex 2 (PRC2). PRC2 regulates gene silencing by catalyzing the methylation of histone H3 at lysine 27. Both overexpression and mutation of EZH2 are associated with the incidence and aggressiveness of various cancers. The novel crystal structure of the SET domain was determined in order to understand disease-associated EZH2 mutations and derive an explanation for its inactivity independent of complex formation. The 2.00 Å crystal structure reveals that, in its uncomplexed form, the EZH2 C-terminus folds back into the active site blocking engagement with substrate. Furthermore, the S-adenosyl-L-methionine (SAM) binding pocket observed in the crystal structure of homologous SET domains is notably absent. This suggests that a conformational change in the EZH2 SET domain, dependent upon complex formation, must take place for cofactor and substrate binding activities to be recapitulated. In addition, the data provide a structural context for clinically significant mutations found in the EZH2 SET domain.

Aluminum concentrations of sand filter and polymeric membrane filtrates: A comparative study

The residual aluminum concentration in treated water after aluminum coagulation and solid–liquid separation is one of the key factors in evaluating the effectiveness of water treatments. Polymeric membrane filters have been widely used to operationally define dissolved substances from particulate matter, and have sometimes been employed in laboratories and pilot studies to approximately simulate media filtration. A comparison between media and membrane filters, however, has not yet been fully investigated. This study compared aluminum concentrations in the filtrate after sand bed and membrane filtrations. The aluminum concentration after sand bed filtration was lower than that after filtration using a membrane with a pore size of 0.1μm, irrespective of the membrane material, with one exception. A PVDF membrane with a pore size of 0.1μm produced a similar residual aluminum concentration to that obtained using a sand bed. When the natural organic matter concentration was very low, the aluminum concentration in the PVDF membrane (pore size 0.1μm) and sand bed filtrates was lower than that obtained using other membranes with the same pore size. It is believed that adsorption of dissolved aluminum may enhance aluminum removal by the sand bed and the PVDF membrane when the aluminum exists in an uncomplexed form with natural organic matter.

Metal-based Antimicrobial Agents: Synthesis, Characterization and Biological Studies of Mannich Base Derivatives of Benzimidazole and Their Metal Complexes

Novel Mannich base derivatives of Benzimidazole were prepared through the condensation reaction of benzimidazole derivative with formaldehyde and primary and/ secondary amine. Zinc(II), copper(II), nickel(II) and cobalt(II) complexes of Mannich bases have also been synthesized. All the compounds were fully characterized by, elemental analyses, magnetic moment determination, molar conductivity measurement, thermogravimetric analysis, spectral and analytical data. Experimental results showed that metal complexes act as bi-dentate ligands towards divalent metal ions via azomethine-N and deprotonated-O while suggesting an octahedral geometry. All the compounds were screened for in-vitro antibacterial and antifungal activity against various bacterial and fungal strains. Almost all the compounds showed good potent activity against microorganisms. It was also seen that compounds with complexed form were more active as compared to un-complexed form. The prepared compounds were also screened for their cytotoxicity and results showed that only Ni(II) complexes exhibit some cytotoxicity while all other compounds were almost inactive.

Specific Conformational States of Ras GTPase upon Effector Binding

To uncover the structural and dynamical determinants involved in the highly specific binding of Ras GTPase to its effectors, the conformational states of Ras in uncomplexed form and complexed to the downstream effectors Byr2, PI3Kγ, PLCε, and RalGDS were investigated using molecular dynamics and cross-comparison of the trajectories. The subtle changes in the dynamics and conformations of Ras upon effector binding require an analysis that targets local changes independent of global motions. Using a structural alphabet, a computational procedure is proposed to quantify local conformational changes. Positions detected by this approach were characterized as either specific for a particular effector, specific for an effector domain type, or as effector unspecific. A set of nine structurally connected residues (Ras residues 5–8, 32–35, 39–42, 55–59, 73–78, and 161–165), which link the effector binding site to the distant C-terminus, changed dynamics upon effector binding, indicating a potential effector-unspecific signaling route within the Ras structure. Additional conformational changes were detected along the N-terminus of the central β-sheet. Besides the Ras residues at the effector interface (e.g., D33, E37, D38, and Y40), which adopt effector-specific local conformations, the binding signal propagates from the interface to distant hot-spot residues, in particular to Y5 and D57. The results of this study reveal possible conformational mechanisms for the stabilization of the active state of Ras upon downstream effector binding and for the structural determinants responsible for effector specificity.