Impact of nanosilver surface electronic distributions on serum protein interactions and hemocompatibility.

The translation of silver-based nanotechnology 'from bench to bedside' requires a deep understanding of the molecular aspects of its biological action, which remains controversial at low concentrations and non-spherical morphologies. Here, we present a hemocompatibility approach based on the effect of the distinctive electronic charge distribution in silver nanoparticles (nanosilver) on blood components. According to spectroscopic, volumetric, microscopic, dynamic light scattering measurements, pro-coagulant activity tests, and cellular inspection, we determine that at extremely low nanosilver concentrations (0.125-2.5µg ml-1), there is a relevant interaction effect on the serum albumin and red blood cells (RBCs). This explanation has its origin in the surface charge distribution of nanosilver particles and their electron-mediated energy transfer mechanism. Prism-shaped nanoparticles, with anisotropic charge distributions, act at the surface level, generating a compaction of the native protein molecule. In contrast, the spherical nanosilver particle, by exhibiting isotropic surface charge, generates a polar environment comparable to the solvent. Both morphologies induce aggregation at NPs/bovine serum albumin ≈ 0.044 molar ratio values without altering the coagulation cascade tests; however, the spherical-shaped nanosilver exerts a negative impact on RBCs. Overall, our results suggest that the electron distributions of nanosilver particles, even at extremely low concentrations, are a critical factor influencing the molecular structure of blood proteins' and RBCs' membranes. Isotropic forms of nanosilver should be considered with caution, as they are not always the least harmful.

Exploring the BSA- and DNA-binding, cytotoxicity, and cell cycle evaluation of ternary copper(II)/diimine complexes with N,N-dibenzyl-N'-benzoylthiourea as promising metallodrug candidates.

Four new copper(II) complexes were synthesized and characterized with the general formula [Cu(N-N)(Th)(NO3)], where N-N corresponds to the N-heterocyclic ligands 1,10-phenanthroline (phen), 2,2'-bipyridine (bipy), 4,7-diphenyl-1,10-phenanthroline (dpp), and 4,4-dimethyl-2,2'-bipyridine (dmbp) and Th represents the N,N-dibenzyl-N'-benzoylthiourea. Cytotoxic activities of the complexes against HCT116 (human colon carcinoma), HepG2 (human hepatocellular carcinoma), and non-tumor MRC-5 (human lung fibroblast) cells were investigated. The copper(II) complexes 1-4 were characterized by spectroscopic techniques while complexes 1 and 2 were studied using single-crystal X-ray diffraction as well. The complexes possessed a five-coordinated structure with one nitrate ligand as a monodentate at the axial position and two bidentate ligands N-heterocyclic and N,N-dibenzyl-N'-benzoylthiourea. The complexes showed promising IC50 values, ranging from 0.3 to 9.0 µM. Furthermore, interaction studies with biomolecules such as calf thymus DNA (ct-DNA) and Bovine Serum Albumin (BSA), which can act as possible biological targets of the complexes, were carried out. The studies suggested that the compounds interact moderately with ct-DNA and BSA. Complexes 1, 2, and 4 did not lead to cell accumulation at any stage of the cell cycle but caused a significant increase in internucleosomal DNA fragmentation. Whereas, compound 3 caused cell cycle arrest in the S phase while doxorubicin caused cell cycle arrest in the G2/M phase. The effect of structural modifications on the metal compounds was correlated with their biological properties and it was concluded that an increase in biological activity occurred with increasing the extension of the diimine ligands. Thus, complex 3 was the most promising one.

Molecular dynamics of structural effects of reactive carbonyl species derivate of lipid peroxidation on bovine serum albumin.

BACKGROUND: Serum albumin is the most abundant protein in the Mammalia blood plasma at where plays a decisive role in the transport wide variety of hydrophobic ligands. BSA undergoes oxidative modifications like the carbonylation by the reactive carbonyl species (RCSs) 4-hydroxy-2-nonenal (HNE), 4 hydroxy-2-hexenal (HHE), malondialdehyde (MDA) and 4-oxo-2-nonenal (ONE), among others. The structural and functional changes induced by protein carbonylation have been associated with the advancement of neurodegenerative, cardiovascular, metabolic and cancer diseases. METHODS: To elucidate structural effects of protein carbonylation with RCSs on BSA, parameters for six new non-standard amino acids were designated and molecular dynamics simulations of its mono­carbonylated-BSA systems were conducted in the AMBER force field. Trajectories were evaluated by RMSD, RMSF, PCA, RoG and SASA analysis. RESULTS: An increase in the conformational instability for all proteins modified with local changes were observed, without significant changes on the BSA global three-dimensional folding. A more relaxed compaction level and major solvent accessible surface area for modified systems was found. Four regions of high molecular fluctuation were identified in all modified systems, being the subdomains IA and IIIB those with the most remarkable local conformational changes. Regarding essential modes of domain movements, it was evidenced that the most representatives were those related to IA subdomain, while IIIB subdomain presented discrete changes. CONCLUSIONS: RCSs induces local structural changes on mono­carbonylated BSA. Also, this study extends our knowledge on how carbonylation by RCSs induce structural effects on proteins.

Role for Carboxylic Acid Moiety in NSAIDs: Favoring the Binding at Site II of Bovine Serum Albumin.

The molecular structures of nonsteroidal anti-inflammatory drugs (NSAIDs) vary, but most contain a carboxylic acid functional group (RCOOH). This functional group is known to be related to the mechanism of cyclooxygenase inhibition and also causes side effects, such as gastrointestinal bleeding. This study proposes a new role for RCOOH in NSAIDs: facilitating the interaction at the binding site II of serum albumins. We used bovine serum albumin (BSA) as a model to investigate the interactions with ligands at site II. Using dansyl-proline (DP) as a fluorescent site II marker, we demonstrated that only negatively charged NSAIDs such as ibuprofen (IBP), naproxen (NPX), diflunisal (DFS), and ketoprofen (KTP) can efficiently displace DP from the albumin binding site. We confirmed the importance of RCOO by neutralizing IBP and NPX through esterification, which reduced the displacement of DP. The competition was also monitored by stopped-flow experiments. While IBP and NPX displaced DP in less than 1 s, the ester derivatives were ineffective. We also observed a higher affinity of negatively charged NSAIDs using DFS as a probe and ultrafiltration experiments. Molecular docking simulations showed an essential salt bridge between the positively charged residues Arg409 and Lys413 with RCOO-, consistent with the experimental findings. We performed a ligand dissociation pathway and corresponding energy analysis by applying molecular dynamics. The dissociation of NPX showed a higher free energy barrier than its ester. Apart from BSA, we conducted some experimental studies with human serum albumin, and similar results were obtained, suggesting a general effect for other mammalian serum albumins. Our findings support that the RCOOH moiety affects not only the mechanism of action and side effects but also the pharmacokinetics of NSAIDs.

Fluorescence study of the interaction between albumin and layered double hydroxides.

Layered double hydroxides nanoparticles (LDH-NP) are increasingly studied for biomedical applications. Nevertheless, their interaction with biomolecules such as proteins needs further exploration for an effective application. In this work, the adsorption of bovine serum albumin (BSA) on LDH-NP and the conformation changes of the protein upon adsorption were characterized using fluorescence spectroscopy. First, the quenching of tryptophan residues of BSA by chloride-intercalated LDH-NP was explored and the BSA adsorption capacity of LDH-NP were determined. Then, the structural conformation of the protein was analyzed by fluorescence spectroscopy (including synchronous, polarization and quenching studies) at different surface coverages. Finally, the proclivity of adsorbed BSA molecules to assemble as amyloid fibril was evaluated. Due to the positive charging and low curvature of LDH-NP, BSA molecules were strongly adsorbed, which produced a quenching of the protein fluorescence and a large adsorption capacity. The effect on BSA conformation was dependent on surface coverage (SC): at low values ,t he tryptophan residues were in more hydrophobic environments and more accessible to quenchers than al high ones. At low SC, there is space between the BSA molecules to spread on the surface, which led to a conformation change. Contrarily, the native conformation around tryptophan residues of BSA was preserved at high SC due to the tight packing of the adsorbed protein molecules. As a result, BSA molecules are stabilized against the formation of amyloid fibrils at high SC, while at low SC they present a similar fibrillation than free BSA.

Radioactive gold nanoparticles coated with BSA: A promising approach for prostate cancer treatment.

Background: Nanotechnology has revolutionized medicine, especially in oncological treatments. Gold nanoparticles (AuNPs) stand out as an innovative alternative due to their biocompatibility, potential for surface modification, and effectiveness in radiotherapeutic techniques. Given that prostate cancer ranks as one of the leading malignancies among men, there's a pressing need to investigate new therapeutic approaches. Methods: AuNPs coated with bovine serum albumin (BSA) were synthesized and their cytotoxicity was assessed against prostate tumor cell lines (LNCaP and PC-3), healthy prostate cells (RWPE-1), and endothelial control cells (HUVEC) using the MTS/PMS assay. For in vivo studies, BALB/C Nude mice were employed to gauge the therapeutic efficacy, biodistribution, and hematological implications post-treatment with BSA-coated AuNPs. Results: The BSA-coated AuNPs exhibited cytotoxic potential against PC-3 and LNCaP lines, while interactions with RWPE-1 and HUVEC remain subjects for further scrutiny. Within animal models, a diverse therapeutic response was observed, with certain instances indicating complete tumor regression. Biodistribution data emphasized the nanoparticles' affinity towards particular organs, and the majority of hematological indicators aligned with normative standards. Conclusions: BSA-coated AuNPs manifest substantial promise as therapeutic tools in treating prostate cancer. The present research not only accentuates the nanoparticles' efficacy but also stresses the imperative of optimization to ascertain both selectivity and safety. Such findings illuminate a promising trajectory for avant-garde therapeutic modalities, holding substantial implications for public health advancements.

New ternary Fe(III)-8-hydroxyquinoline-reduced Schiff base complexes as selective anticancer drug candidates.

Due to the growing prevalence of cancer diseases, new therapeutic options are urgently needed, and drugs based on metal ions other than platinum are alternatives with exciting possibilities. We report the synthesis, characterization and biological effect of mixed-ligand Fe(III)-aminophenolate complexes derived from salicylaldehyde and L-tryptophan with quinoline derivatives as co-ligands, namely 8-hydroxyquinoline (8HQ), [Fe(L)(8HQ)(H2O)] (1) and its 5-cloro derivative (Cl8HQ), [Fe(L)(Cl8HQ)(H2O)] (2). The complex bearing the aminophenolate and lacking the quinoline co-ligand, [Fe(L)(Cl)(H2O)2] (3), was prepared for comparison. The analytical and spectroscopic characterization revealed that 1 and 2 are octahedral Fe(III) complexes with the aminophenolate acting as a dianionic tridentate ligand and 8HQ co-ligands as bidentate chelates. Spectroscopic techniques and molecular docking studies were used to evaluate the ability of these complexes to bind bovine serum albumin (BSA) and calf thymus DNA. Complex 2 [Fe(L)(Cl8HQ)(H2O)] was the one showing higher affinity for both biomolecules. Cell viability was assessed in breast, colorectal and bone human cancer cell lines. 1 and 2 were found to be more active than cisplatin in all cell lines tested. A non-tumoral fibroblast line (L929, mouse non-tumoral fibroblasts) was used to evaluate selectivity. The results evidence that 2 shows much higher selectivity than 1 in all cell lines tested, but particularly in bone cancer cells in which selectivity index (SI) values are 8.0 and 18.8 for 1 and 2, respectively.

Amphotericin B resistance in Leishmania mexicana: Alterations to sterol metabolism and oxidative stress response.

Amphotericin B is increasingly used in treatment of leishmaniasis. Here, fourteen independent lines of Leishmania mexicana and one L. infantum line were selected for resistance to either amphotericin B or the related polyene antimicrobial, nystatin. Sterol profiling revealed that, in each resistant line, the predominant wild-type sterol, ergosta-5,7,24-trienol, was replaced by other sterol intermediates. Broadly, two different profiles emerged among the resistant lines. Whole genome sequencing then showed that these distinct profiles were due either to mutations in the sterol methyl transferase (C24SMT) gene locus or the sterol C5 desaturase (C5DS) gene. In three lines an additional deletion of the miltefosine transporter gene was found. Differences in sensitivity to amphotericin B were apparent, depending on whether cells were grown in HOMEM, supplemented with foetal bovine serum, or a serum free defined medium (DM). Metabolomic analysis after exposure to AmB showed that a large increase in glucose flux via the pentose phosphate pathway preceded cell death in cells sustained in HOMEM but not DM, indicating the oxidative stress was more significantly induced under HOMEM conditions. Several of the lines were tested for their ability to infect macrophages and replicate as amastigote forms, alongside their ability to establish infections in mice. While several AmB resistant lines showed reduced virulence, at least two lines displayed heightened virulence in mice whilst retaining their resistance phenotype, emphasising the risks of resistance emerging to this critical drug.

Toxicity mitigation and biodistribution of albumin corona coated graphene oxide and carbon nanotubes in Caenorhabditis elegans.

In this work, the toxicity and biodistribution of graphene oxide (GO) and oxidized multi-walled carbon nanotubes (MWCNT) were investigated in Caenorhabditis elegans. Bovine serum albumin (BSA) was selected as a model protein to evaluate the influence of protein corona formation on materials physicochemical properties, colloidal stability, and toxicity. Biological assays were performed to assess the effects of bare and albumin corona coated materials on survival, oxidative stress, intestinal barrier permeability, growth, reproduction, and fertility. Critical alterations in topography, surface roughness and chemistry of GO and MWCNT were observed due to albumin corona formation. These modifications were associated with changes in colloidal stability of materials and prevention of their aggregation and sedimentation in nematode testing medium. Both GO and MWCNT caused damage to nematode survival, growth, reproduction, and fertility, as well as enhanced oxidative stress and permeability of the intestinal barrier. But GO was more toxic than MWCNT to C. elegans, especially at long-term assays. Albumin corona mitigated 100% of acute and chronic effects of MWCNT. In contrast, the negative effects of GO were not completely mitigated; GO inhibited 16.2% of nematode growth, 86.5% of reproduction, and 32.0% of fertility at the highest concentration evaluated (10 mg L-1), while corona coated GO mitigated 50% and 100% of fertility and growth, respectively. Confocal Raman spectroscopy imaging was crucial to point out that bare and albumin corona coated GO and MWCNT crossed the C. elegans intestinal barrier reaching its reproductive organs. However, BSA corona protected the nematodes targeted organs from negative effects from MWCNT and blocked its translocation to other tissues, while coated GO was translocated inside the nematode affecting the functionality of crucial organs. In addition, coated MWCNT was excreted after 2 h of food resumption, whereas coated GO still accumulated in the nematode intestine. Our results demonstrate that the materials different translocation and excretion patterns in C. elegans had a relation to the impaired physiological functions of primary and secondary organs. This work is a contribution towards a better understanding of the impacts of protein corona on the toxicity of graphene oxide and carbon nanotubes; essential information for biological applications and nanosafety.

The proteomic and particle composition of human platelet lysate for cell therapy products.

Following health agencies warning, the use of animal origin supplements should be avoided in biological products proposed as therapy in humans. Platelet lysate and several other growth factors sources are alternatives to replace fetal calf serum, the current gold standard in clinical-grade cell culture. However, the platelet supplement's content lacks data due to different production methods. The principle behind these products relays on the lysis of platelets that release several proteins, some of which are contained in heterogeneous granules and coordinate biological functions. This study aims to analyze the composition and reproducibility of a platelet lysate produced with a standardized method, by describing several batches' protein and particle content using proteomics and dynamic light scattering. Proteomics data revealed a diversified protein content, with some related to essential cellular processes such as proliferation, morphogenesis, differentiation, biosynthesis, adhesion, and metabolism. It also detected proteins responsible for activation and binding of transforming growth factor beta, hepatocyte growth factor, and insulin-like growth factor. Total protein, biochemical, and growth factors quantitative data showed consistent and reproducible values across batches. Novel data on two major particle populations is presented, with high dispersion level at 231 ± 96 d.nm and at 30 ± 8 d.nm, possibly being an important way of protein trafficking through the cellular microenvironment. This experimental and descriptive analysis aims to support the content definition and quality criteria of a cell supplement for clinical applications.

Study on the cytotoxic, antimetastatic and albumin binding properties of the oxidovanadium(IV) chrysin complex. Structural elucidation by computational methodologies.

We have previously synthesized and characterized the chrysin coordination complex with the oxidovanadium(IV) cation (VIVO(chrys)2) and characterized in ethanolic solution and in solid state. Because suitable single crystals for X-ray diffraction determinations could not be obtained, in the present work, we elucidate the geometrical parameters of this complex by computational methodologies. The optimization and vibrational investigation were carried out both in ethanolic solution and in gas phase. The computational results support the experimentally proposed geometries of the VIVO(chrys)2 complex, thus leading to the conclusion that the complex exists as conformers with trans-octahedral geometry in ethanolic solution and as conformers with cis-octahedral geometry in the solid state. The complex also exists as conformers with trans-octahedral geometry in aqueous media. The active species formed after dissolution in DMSO showed anticancer and antimetastatic behavior in human lung cell line A549 with moderate binding (Kaca. 105 M-1) to bovine serum albumin (BSA). The interaction through hydrogen bonding and van der Waals forces resulted in a spontaneous process. Site marker competitive experiments showed binding sites for chrysin mainly located in site II (subdomain IIIA) and in site I (subdomain IIIA) for the complex. FT-IR spectral measurements showed evidences of the alterations of protein secondary structure in the presence of chrysin and VIVO(chrys)2.

Candesartan, losartan and valsartan Zn(II) complexes interactions with bovine serum albumin.

Background: The pharmacological response and the therapeutic efficacy of a drug depends on the interactions with plasma proteins. Methodology: The interaction of bovine serum albumin (BSA) with the metal complexes of antihypertensive drugs, Zn(II)/sartan complexes (candesartan, valsartan and losartan), was investigated using fluorescence quenching determinations at different temperatures. Results: The binding studies of the compounds with BSA showed static quenching and moderate binding with calculated constants in the range of 104-106 M-1, indicating potent serum distribution via albumins. In all cases, negative values of free energy are indicative of spontaneous processes and the stabilization of BSA/compound complexes through hydrogen bonding and van der Waals forces. The results for the sartans agree with the reported pharmacokinetics studies. Conclusion: It has been determined that the three sartans and the Zn complexes could be transported and distributed by albumin.

Molecular insight on the binding of stevia glycosides to bovine serum albumin.

The interaction of the steviol and its glycosides (SG), steviolbioside, and rebaudioside A, with bovine serum albumin (BSA) was studied by absorption and fluorescence spectroscopy techniques alongside molecular docking. The stevia derivatives quenched the fluorescence of BSA by a dynamic quenching mechanism, indicating the interaction between the stevia derivatives and BSA. The binding constant (Kb) of steviol was 100-1000-fold higher than those of SG. The stevia derivative/BSA binding reaction was spontaneous and involved the formation of hydrogen bonds and van der Waals interactions between steviol and steviolbioside with BSA, and water reorganization around the rebaudioside A/BSA complex. Molecular docking pointed out the FA1 and FA9 binding sites of BSA as the probable binding sites of steviol and SG, respectively. In conclusion, steviol enhanced hydrophobicity and small size compared to SG may favor its binding to BSA. As steviol and its glycosides share binding sites on BSA with free fatty acids and drugs, they may be competitively displaced from plasma albumin under various physiological states or disease conditions. These findings are clinically relevant and provide an insight into the pharmacokinetics and pharmacodynamics of the stevia glycosides.

A novel ruthenium(ii) gallic acid complex disrupts the actin cytoskeleton and inhibits migration, invasion and adhesion of triple negative breast tumor cells.

This work describes the synthesis of three new ruthenium(ii) complexes with gallic acid and derivatives of the general formula [Ru(L)(dppb)(bipy)]PF6, where L = gallate (GAC), benzoate (BAC), and esterified-gallate (EGA), bipy = 2,2'-bipyridine and dppb = 1,4-bis(diphenylphosphino)butane. The complexes were characterized by elemental analysis, molar conductivity, NMR, cyclic voltammetry, UV-vis and IR spectroscopy, and two of them by X-ray crystallography. Cell viability assays show promising results, indicating higher cytotoxicity of the complexes in MDA-MB-231 cells, a triple-negative breast cancer (TNBC) cell line, compared with the hormone-dependent MCF-7 cell line. Studies in vitro with the MDA-MB-231 cell line showed that only Ru(BAC) and Ru(GAC) interacted with BSA. Besides that, the Ru(GAC) complex, which has a polyphenolic acid, interacted in an apo-Tf structure and function dependent manner and it was able to inhibit the formation of reactive oxygen species. Ru(GAC) was able to cause damage to the cellular cytoskeleton leading to inhibition of some cellular processes of TNBC cells, such as invasion, migration, and adhesion.

Effects of the age/rage axis in the platelet activation.

Platelet activity is essential in cardiovascular diseases. Therefore our objective was to evaluate the main effects of activating RAGE in platelets which are still unknown. A search for RAGE expression in different databases showed poor or a nonexistent presence in platelets. We confirmed the expression in platelets and secreted variable of RAGE (sRAGE). Platelets from elderly adults expressed in resting showed 3.2 fold more RAGE from young individuals (p < 0.01) and 3.3 fold with TRAP-6 (p < 0.001). These results could indicate that the expression of RAGE is more inducible in older adults. Then we found that activating RAGE with AGE-BSA-derived from methylglyoxal and subthreshold TRAP-6, showed a considerable increase with respect to the control in platelet aggregation and expression of P-selectin (respectively, p < 0.01). This effect was almost completely blocked by using a specific RAGE inhibitor (FSP-ZM1), confirming that RAGE is important for the function and activation platelet. Finally, we predict the region stimulated by AGE-BSA is located in region V of RAGE and 13 amino acids are critical for its binding. In conclusion, the activation of RAGE affects platelet activation and 13 amino acids are critical for its stimulation, this information is crucial for future possible treatments for CVD.

Modified pyrazole platinum(II) complex can circumvent albumin and glutathione: Synthesis, structure and cytotoxic activity.

The synthesis and structural characterization of novel platinum complexes ([PtII(Pz)2Cl2] - C1, C2 and C3) featuring diphenyl-pyrazole derived ligands: para-fluorophenyl and para-substituted phenyl (CH3, F and Cl for L1, L2 and L3, respectively) were reported and it was also evaluated their potential antitumor activity. The elemental, molar conductivity and thermogravimetric analysis combined with FTIR, UV-vis, NMR and mass spectrometry are in agreement with the chemical structure indicated by single-crystal X-ray diffraction. The antiproliferative activities were assessed against tumor (B16F10 and 4T1) and non-tumor (BHK21) cell lines, and the cytotoxicity of the compounds was strongly increased after metal complexation displaying promising activity. It was also assessed the ability of extracellular bovine serum albumin (BSA) and glutathione (GSH) to decrease the cytotoxicity of the complexes against B16F10. It was highlighted that only the C3 activity was not disturbed in those conditions, being confirmed by flow cytometry using Anexin-V/PI to evaluate interferences in the apoptosis process, even it was not predicted by molecular docking simulations. The interaction of the synthesized compounds with calf-thymus DNA (ctDNA) and bovine serum albumin (BSA) was also investigated through spectrophotometric assays and molecular docking simulations, indicating that C1 and C2 presented better interaction with the biomacromolecules than the corresponding ligands. In addition, agarose gel electrophoresis with plasmid DNA revealed that C1-C3 are capable of interaction with DNA and modify its electrophoretic mobility.

Avenanthramide-C prevents amyloid formation of bovine serum albumin.

The misfolding of protein and its assembly into amyloid fibrils with a characteristic ß-sheet-rich secondary structure, cause a lot of illnesses. Polyphenols have been extensively studied as a class of amyloid inhibitors, whose effect depends on the position and number of hydroxyl groups around the flavone backbone. In this study, we used bovine serum albumin (BSA) as an amyloid model to test the anti-amyloid effects of Avenanthramide-C (Avn-C), a molecule with a long aliphatic linker between two aromatic rings. We used spectroscopy techniques like thioflavin T fluorescence and circular dichroism, to follow the ß-sheet-rich aggregates of BSA upon incubation at 68 °C. Our results demonstrated that Avn-C shows higher inhibitory effect on BSA oligomerization at micromolar concentrations, than Epigallocatechin gallate (EGCG) and Curcumin, proving for the first time, that Avn-C can serve as potential molecule in preventing protein aggregation.

A novel Cu(II) distorted cubane complex containing Cu4O4 core as the first tetranuclear catalyst for temperature dependent oxidation of 3,5-di-tert-butyl catechol and in interaction with DNA & protein (BSA).

The tri-dentate Schiff base ligand 3-(2-hydroxyethylimino)-1-phenylbut-1-en-1-ol (L) produced the tetra-nuclear Cu(II) distorted cubane complex which contain Cu4O4 core, upon reaction with Cu(II)acetate.H2O. The complex was structurally characterized by X-ray crystallography and found that, in this tetrameric and tetra-nuclear distorted cubane structure, each two-fold deprotonated Schiff base ligand coordinated to a Cu(II) center with their alcoholic oxygens and imine nitrogens and formed six and five-membered chelate rings. At the same time, each ligand bridged to a neighboring Cu(II) atom by its alcoholic oxygen, thus the metal centers became penta-coordinated. The copper(II) complex with µ-ɳ2-hydroxo bridges and Cu….Cu distance about 3 Šwas structurally similar to the active site of natural catechol oxidase enzyme and exhibited excellent catecholase activity in aerobic oxidation of 3,5-di-tert-butyl catechol to its o-quinone. The kinetics and mechanism of the oxidation of 3, 5-DTBCH2 catalyzed by [CuL]4 complex, were studied at four different temperatures from 283 to 313K by UV-Vis spectroscopy. Interaction of [CuL]4 complex with FS-DNA was investigated by UV-Vis and fluorescence spectroscopy, viscosity measurements, cyclic voltammetry (CV), circular dichroism (CD) and agarose gel electrophoresis. The main mode of binding of the complexes with DNA was intercalation. The interaction between [CuL]4 complex and bovine serum albumin (BSA) was studied by UV-Vis, fluorescence and synchronous fluorescence spectroscopic techniques. The results indicated a high binding affinity of the complex to BSA. In vitro anticancer activity of the complex was evaluated against A549, Jurkat and Ragi cell lines by MTT assay. The complex was remarkably active against the cell lines and can be a good candidate for an anticancer drug. Theoretical docking studies were performed to further investigate the DNA and BSA binding interactions.

B procyanidins of Annona crassiflora fruit peel inhibited glycation, lipid peroxidation and protein-bound carbonyls, with protective effects on glycated catalase.

Advanced glycation end-products (AGEs) have been reported as results of increased oxidative stress. Consequently, the search for new antioxidant and anti-glycating agents is under intense investigation. Plant-derived procyanidins have previously demonstrated anti-glycation properties. Thus, this study aimed to isolate procyanidins from Annona crassiflora fruit peel, a species from the Brazilian Savanna, and investigate their antioxidant and anti-glycation effects. Free radical scavenging and quenching properties, formation of reactive oxygen species (ROS), AGEs, protein carbonyl and thiol groups, lipid peroxidation, crosslinked AGEs, as well as glycated catalase activity, were analyzed. In addition, in silico assessment of absorption, distribution, metabolism, excretion and toxicity was carried out. The procyanidins-enriched fraction, named here as F7, showed high antioxidant and anti-glycation capacities, with inhibitory activities against lipid peroxidation, and AGEs and ROS formation. In addition, there were reductions in AGEs-induced crosslinks and protein carbonyls and protective effects against oxidation of thiol groups and glycated-catalase. ADMET predictions of F7 showed favorable absorption and distribution, with no hepatotoxicity or mutagenicity. Together, our results support the anti-glycation activities of the procyanidins-enriched fraction from A. crassiflora, and suggest that these effects are triggered, at least in part, by scavenging free radical and dicarbonyls intermediates.

High levels of serum glycans monovalent IgG immune complexes detected by dissociative ELISA in experimental visceral leishmaniasis.

Visceral leishmaniasis (VL) is epidemic in Brazil with an increasing incidence of human cases and canine reservoirs, with host hypergammaglobulinemia. Conventional enzyme-linked immunosorbent assay (cELISA) based on several parasitic antigens is the main method for diagnosis and indication of treatment. Dissociative ELISA (dELISA) uses acidic treatment to free immunoglobulin G (IgG) from immune complexes, and its use revealed a significant positive fraction of suspected cases with negative serology. Looking for small molecules or haptens that block IgG antibodies, we purified by molecular exclusion chromatography, 1000-3000 MW molecules from promastigote soluble extract, mostly oligosaccharides comprising 6-13 sugar residues using MALDI-TOF analysis. Glycan-BSA complex (GBC) was constructed by conjugating promastigote glycans to BSA molecules, allowing their use in the solid support in cELISA or dELISA. Sera from experimentally infected hamsters showed higher levels of blocked monomeric IgG during infection, mostly against GBC, which was also present in lower concentrations in the promastigote soluble extract dELISA. Those data show that most of the specific monomeric IgG in serum are blocked by haptens composed by glycans produced by the parasite, better detected in the high dilution of sera in the dELISA assays. dELISA is a useful technique for detecting blocked monomeric antibodies that could have difficult clearance from blood, which could result in hypergammaglobulinemia.