Search

Bio-oil derived from the pyrolysis of lignocellulosic biomass residues cannot be used directly as biofuel due to the high content of oxygenated compounds. As an alternative, bio-oil must undergo a deoxygenation process, such as catalytic hydrodeoxygenation (HDO). In this sense, this work studied the effect of different concentrations of Ce3+ and Ce4+ in layered double hydroxides (LDHs) in order to obtain mixed oxide catalysts containing NiAlCe (NiO-NiAl2O4-CeO2) with low cost and high performance for the hydrodeoxygenation of anisole as a model bio-oil compound. Mixed oxides were obtained from the thermal decomposition of layered double hydroxides (LDHs) by using terephthalic acid as compensation anion, with molar ratio: Ni2+/(Al3++ M) = 1.0, where M = Ce3+; Ce4+ or Ce3+-Ce4+; and Al/Ce ratios of 9 and 1. Characterization analyses confirmed the formation of LDHs for all materials, although at lower Al/Ce ratio it is observed a loss of crystallinity, due to a greater repulsion and distortion of the layer structure caused by the incorporation, in greater amount, of cations with a high ionic radius. The increase in the content of cerium in mixed oxides also led to an increase in the acidity of the catalysts, in addition to a reduction in the surface area, justified by the pore blockage by CeO2 on the catalyst surface. The greater structural and thermal stability was evidenced in the LDHs derived from Ce4+, as well as a greater dispersion of the NiO phase in the corresponding mixed oxides, and therefore presenting greater anisole conversion. The obtained data indicated that the presence of Ce4+ ions on the catalyst surface was decisive in the conversion of anisole to cyclohexane, the main deoxygenated product, obtained via the direct deoxygenation and hydrogenation mechanism.

Background Site-specific multimodal nanoplatforms with fluorescent-magnetic properties have great potential for biological sciences. For this reason, we developed a multimodal nanoprobe (BNPs-Tf), by covalently conjugating an optical-magnetically active bimodal nanosystem, based on quantum dots and iron oxide nanoparticles, with the human holo-transferrin (Tf). Methods The Tf bioconjugation efficiency was evaluated by the fluorescence microplate assay (FMA) and the amount of Tf immobilized on BNPs was quantified by fluorescence spectroscopy. Moreover, relaxometric and fluorescent properties of the BNPs-Tf were evaluated, as well as its ability to label specifically HeLa cells. Cytotoxicity was also performed by Alamar Blue assay. Results The FMA confirmed an efficient bioconjugation and the fluorescence spectroscopy analysis indicated that 98% of Tf was immobilized on BNPs. BNPs-Tf also presented a bright fluorescence and a transversal/longitudinal relaxivities ratio (r2/r1) of 65. Importantly, the developed BNPs-Tf were able to label, efficiently and specifically, the Tf receptors in HeLa cells, as shown by fluorescence and magnetic resonance imaging assays. Moreover, this multimodal system did not cause noteworthy cytotoxicity. Conclusions The prepared BNPs-Tf hold great promise as an effective and specific multimodal, highly fluorescent-magnetic, nanoplatform for fluorescence analyses and T2-weighted images. General significance This study developed an attractive and versatile multimodal nanoplatform that has potential to be applied in a variety of in vitro and in vivo studies, addressing biological processes, diagnostic, and therapeutics. Moreover, this work opens new possibilities for designing other efficient multimodal nanosystems, considering other biomolecules in their composition able to provide them important functional properties.

Short chain sodium polyphosphates – (NaPO3)9 – PolyP – were added to 2-in-1 pediatric parenteral nutrition (PN) formulations as new alternative phosphate sources. Solutions of PolyP and Ca2+ in the presence of Mg2+, glucose, amino acids and trace elements were analyzed by electronic absorption and 31P NMR spectroscopies, as well as by pH measurements. Additionally, to verify the biocompatibility of short chain PolyP solutions, red blood cell (RBCs) units were evaluated by analyzing hemoglobin, hematocrit, and hemolysis. The blood clotting action of the PolyP was determined through Prothrombin time. We observed that the short chain PolyP anions did not interfere in the quality control parameters for RBCs units or in the blood extrinsic coagulation pathway. The spectral changes observed in the 31P NMR confirm the association of Mg2+ and Ca2+ with PolyP in the admixtures. Compatibility diagrams showed that the PolyP-Ca/Mg systems reflect a gel behavior based on the PolyP chains interaction with the incoming cations maintaining a supramolecular equilibrium until a limiting concentration. We observed that the greater the PolyP concentration, the greater the amount of Ca2+ that could be added to the admixture without phase separation and that the increased complexity of species (glucose, amino acid, trace elements and Mg2+) in the admixture favored a still higher final Ca2+ concentration. The overall colloidal behavior of short chain PolyPs shows that these polyanions function not only as alternative and safe phosphate sources in PN formulations but also as polyanions that increase the admixture stability against phase separation by efficiently sequestering metal cations present.

Mastitis is a bacterial infection that affects all lactating mammals, and in dairy cattle, it leads to a reduction in their milk production and, in worse cases, it may lead to animal death. One viable therapeutic modality for overcoming bacterial resistance can be photodynamic inactivation (PDI), a therapeutic modality for bacterial infection treatment. One of the main factors that can lead to an efficient PDI process is the association of metallic nanoparticles in the close vicinity of photosensitizers, which has shown promising results due to localized surface plasmon resonance phenomena. In this work, methylene blue (MB) molecules were associated with Ag prismatic nanoplatelets (AgNPrs) to use as PDI photosensitizer against Staphylococcus aureus isolated from bubaline mastitis. The optical plasmonic activity of AgNPrs was tuned to the MB absorption region (600-700 nm) by inducing their growth into prismatic shapes by a seed-mediated procedure, using poly (sodium 4-styrene sulfonate) as the surfactant. A simulation on the plasmonic properties of the nanoprisms, applying particle size within the dimensions determined by TEM image analysis (d = 32 ± 6 nm), showed a 30 % increase of the incident field on the prismatic tips. Photodynamic results showed that the electrostatic AgNPr-MB conjugates promoted enhancement (ca. 15 %) of the reactive oxygen species production. Besides, PDI mediated by AgNPrs-MB led to the complete inactivation of the mastitis S. aureus strain after 6 min inactivation, in contrast to PDI mediated by MB, which reduced less than a 0.5 bacterial log. Thus, the results show this plasmonic enhanced photodynamic tool's potential to be applied in the inactivation of multi-resistant bacterial strains.

Enhancement of hydrophilicity and functionalization of CdTe QDs (Quantum Dots) via surface modifications have made them suitable to be used as specific probes for cell imaging. Applications for targeting cell surfaces have been widely demonstrated in vitro but their use in animal models is not trivial. Here, we reported the interaction of mercaptosuccinic-coated (MSA) CdTe QDs with the epidermis of living and Carnoy-fixed zebrafish embryos. QDs concentrate along adherent junctions and reveal the characteristic pattern of actin microridges at the apical surface of the enveloping layer. In our study, labeling with anionic QDs is attained within few minutes at submicromolar concentrations in whole mounted Carnoy-fixed zebrafish embryos, providing a faster approach compared with immunodetection or standard Phalloidin staining of actin for visualization by fluorescence microscopy. Fil: Sigot, Valeria. Universidad Nacional de Entre Ríos. Instituto de Investigación y Desarrollo en Bioingeniería y Bioinformática - Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Investigación y Desarrollo en Bioingeniería y Bioinformática; Argentina. Universidad Nacional de Entre Ríos. Facultad de Ingeniería; Argentina Fil: Cabral Filho, Paulo E.. Universidade Federal de Pernambuco; Brasil Fil: Sampedro, Maria Florencia. Universidad Nacional de Entre Ríos. Instituto de Investigación y Desarrollo en Bioingeniería y Bioinformática - Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Investigación y Desarrollo en Bioingeniería y Bioinformática; Argentina. Universidad Nacional de Entre Ríos. Facultad de Ingeniería; Argentina Fil: Santos, Beate S.. Universidade Federal de Pernambuco; Brasil Fil: Fontes, Adriana. Universidade Federal de Pernambuco; Brasil

Sialic acids (SAs) modulate essential physiological and pathological conditions, including cell-cell communication, immune response, neurological disorders, and cancer. Besides, SAs confer negative charges to cell membranes, also contributing to hemorheology. Phenylboronic acids, called as mimetic lectins, have been highlighted to study SA profiles. The association of these interesting molecules with the optical properties of quantum dots (QDs) can provide a deeper/complementary understanding of mechanisms involving SAs. Herein, we explored the thiol affinity to the QD surface to develop a simple, fast and direct attachment procedure to functionalize these nanocrystals with 3-mercaptophenylboronic acids (MPBAs). The functionalization was confirmed by fluorescence correlation spectroscopy and inductively coupled plasma spectrometry. The conjugate specificity/efficiency was proved in experiments using red blood cells (RBCs). A labeling >90% was found for RBCs incubated with conjugates, which reduced to 17% after neuraminidase pretreatment. Moreover, QDs-MPBA conjugates were applied in a comparative study using acute (KG-1) and chronic (K562) myelogenous leukemia cell lines. Results indicated that KG-1 membranes have a greater level of SA, with 100% of cells labeled and a median of fluorescence intensity of ca. 2.5-fold higher when compared to K562 (94%). Therefore, this novel QDs-MPBA conjugate can be considered a promising nanoplatform to evaluate SA contents in a variety of biological systems.

Folic acid content was evaluated in food preparations containing wheat and corn flour submitted to baking, deep-frying, and steaming. Commercially fortified flours showed the absence of folic acid. Flours with laboratory folic acid fortification showed 487 and 474 μg of folic acid in 100 g of wheat and corn flours, respectively. In the corn flour preparations, the cake had the highest retention (99%) when compared to couscous (97%). Besides, the cake showed higher retention when compared to the wheat flour preparations due to the interactions of the folic acid with the hydrophobic amino acids of the Zein, a protein found in corn. In wheat flour preparations, vitamin retention was 87%, 80% and 57% in bread, cake, and White sauce respectively. These findings relate to the change of the physicochemical properties of food components that occurs during mixing and cooking of the ingredients.

The increasing applications of quantum dots (QDs) as optic tools in life science have stimulated researchers to evaluate the effects of these nanoprobes in cell viability using a variety of methods, especially colorimetric ones. One of the most applied tests is the MTT assay. In comparison to MTT, for example, the resazurin-based method has the main advantage of not evaluating the cells directly, thus eliminating false-positive results that may arise from the overlap of the absorbances of the QD with the colorimetric compound. Therefore, herein, we describe the resazurin assay as an alternative, simple, quick, sensitivity, reproducible, and nontoxic test to evaluate the in vitro cell viability after QD exposure. Moreover, this test presents an additional advantage; the cells remain viable for complementary experimental procedures, such as cell migration or adhesion.

The optical properties of quantum dots (QDs) make them useful tools for biology, especially when combined with biomolecules such as lectins. QDs conjugated to lectins can be used as nanoprobes for carbohydrate expression analysis, which can provide valuable information about glycosylation changes related to cancer and pathogenicity of microorganisms, for example. In this study, we evaluated the best strategy to conjugate Cramoll lectin to QDs and used the fluorescent labeling of Candida albicans cells as a proof-of-concept. Cramoll is a mannose/glucose-binding lectin with unique biological properties such as immunomodulatory, antiparasitic, and antitumor activities. We probed covalent coupling and adsorption as conjugation strategies at different pH values. QDs conjugated to Cramoll at pH7.0 showed the best labeling efficiency in the fluorescence microscopy analysis. Moreover, QD-Cramoll conjugates remained brightly fluorescent and preserved identical biological activity according to hemagglutination assays. Flow cytometry revealed that approximately 17% of C. albicans cells were labeled after incubation with covalent conjugates, while approximately 92% of cells were labeled by adsorption conjugates (both at pH7.0). Inhibition assays confirmed QD-Cramoll specificity, which reduced the labeling to at most 3%. Therefore, the conjugates obtained by adsorption (pH7.0) proved to be promising and versatile fluorescent tools for glycobiology.

Background Photodynamic inactivation (PDI) is emerging as a promising alternative for cutaneous leishmaniasis (CL). The chemotherapy currently used presents adverse effects and cases of drug resistance have been reported. ZnTnHex-2-PyP4+ is a porphyrin with a high potential as a photosensitizer (PS) for PDI, due to its photophysical properties, structural stability, and cationic/amphiphilic character that can enhance interaction with cells. This study aimed to investigate the photodynamic effects mediated by ZnTnHex-2-PyP4+ on Leishmania parasites. Methods ZnTnHex-2-PyP4+ stability was evaluated using accelerated solvolysis conditions. The photodynamic action on promastigotes was assessed by (i) viability assays, (ii) mitochondrial membrane potential evaluation, and (iii) morphological analysis. The PS-promastigote interaction was studied. PDI on amastigotes and the cytotoxicity on macrophages were also analyzed. Results ZnTnHex-2-PyP4+, under submicromolar concentration, led to immediate inactivation of more than 95% of promastigotes. PDI promoted intense mitochondrial depolarization, loss of the fusiform shape, and plasma membrane wrinkling in promastigotes. Fluorescence microscopy revealed a punctate PS labeling in the parasite cytoplasm. PDI also led to reductions of ca. 64% in the number of amastigotes/macrophage and 70% in the infection index after a single treatment session. No noteworthy toxicity was observed on mammalian cells. Conclusions ZnTnHex-2-PyP4+ is stable against demetallation and more efficient as PS than the ethyl analogue ZnTE-2-PyP4+, indicating readiness for evaluation in in vivo studies as an alternative approach to CL. General significance This report highlighted promising photodynamic effects mediated by ZnTnHex-2-PyP4+ on Leishmania parasites, opening up perspectives for applications in CL pre-clinical assays and PDI of other microorganisms.

During carcinogenesis, changes in the glycosylation can modulate many biological processes. Thus, the interest in exploring and understanding the roles of carbohydrates as cancer biomarkers has been increasing. Lectins have been applied as useful tools in glycobiology, especially when associated with fluorescent reporters. Therefore, to take advantage of the physicochemical properties of quantum dots (QDs), herein, we conjugated Cramoll, a lectin that recognizes glucose/mannose residues, with those nanoparticles. We applied the conjugates to investigate the glycocode of normal, fibroadenoma (FB), and invasive ductal carcinoma (IDC) human breast tissues. Additionally, we proposed a method to quantitatively evaluate the tissue labeling intensity by a fluorescence microplate assay (FMA). Conjugates showed intense fluorescence and specificity. The lectin activity and secondary structure were also preserved after the conjugation with QDs. Moreover, fluorescence images showed that ductal cells of normal and FB tissues were preferentially labeled by conjugates, whereas both cells and stroma were strongly labeled in IDC. FMA showed in a quantitative, practical, and sensitive way that the level of exposed glucose/mannose residues increased accordingly to the sample malignancy degree. In conclusion, QDs-Cramoll conjugates can be considered effective, specific, and versatile probes to evaluate glycan profiles in normal and transformed tissues, by fluorescence microscopy as well as FMA quantification. Furthermore, FMA showed to be a potential method that can be applied with other fluorescent conjugates.

In this study we evaluated the absolute fluorescence quantum yield ( Φ ) of hydrophilic CdTe QDs in function of different pHs, modified from the alkaline to acid, by using two different chemicals compounds, the mercaptosuccinic acid (MSA-the stabilizing agent of the QDs synthesis) or hydrochloric acid (HCl). The pH control of QDs suspensions is essential for the use of fluorescent nanoparticles in biological systems. We used the eclipsing thermal lens spectroscopy technique to determine the absolute fluorescence quantum yield values. The results showed variations on the Φ values as a function of the pH, which allowed a better understanding of QDs emission characteristics, establishing parameters for their use in biomedical applications such as optical images of biological systems, immunoassays, flow cytometry, biosensors and others.

In this study, we present the optimization of the optical properties of Mn2+ doped ZnSe QDs (also referred to as d-dots) coated with thioglycolic acid in aqueous medium. The nanoparticles were characterized by ionic coupled plasma, electron paramagnetic resonance, transmission electron microscopy and X-ray diffractometry. By applying a controlled dose of UV irradiation, we obtained efficient orange emitting d-dots (4T1 6A1 transition centered at 580 nm). The results point out to a doping fraction in the nanoparticles smaller than 1% (0.89%), and that the Mn2+ ions are preferentially located close to the particle's surface. The UV photoactivation procedure has a definite influence on the emission intensity and on the colloidal stability of the particles. Photoactivated Mn2+ d-dots were conjugated to Concanavalin A and labeled specifically Candida albicans yeast cells, by targeting carbohydrate residues depicting the successful use of these QDs as fluorescent probes. (© 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim)

We developed a biosensing platform by electropolymerizing 2-(1H-Pyrrol-1-yl)ethanamine on glassy carbon (GC) surfaces, followed by the immobilization of carboxyl-coated CdTe QDs onto the films. The immobilization of QDs was studied by applying two procedures: adsorption (electrostatic interactions) and covalent (using crosslinking agents). At a later stage, the covalent immobilization of IgG antibodies was also tested in platforms prepared with and without QDs. The formation of the polymeric film was confirmed by scanning electron microscopy. Furthermore, all surface modifications were monitored using cyclic voltammetry and electrochemical impedance spectroscopy, and K3[Fe(CN)6]/K4[Fe(CN)6] as the redox probe. The covalent crosslinking showed reproducibility and effective QD immobilization. Our study also showed that the QD coating by carboxylic groups leads to a decrease in the charge transfer process. The nanometric size of QDs and their active surfaces highly increased the functional platform area, enabling a high number of available sites for IgG antibodies. Indeed, in the absence of QDs, the platform was not able to detect the anti-IgG molecules. These results were supported by saturation curves constructed from differential pulse voltammetry. Thus, the use of carboxyl-coated CdTe QDs combined with 2-(1H-Pyrrol-1-yl)ethanamine polymer favored the development of a promising and sensitive biosensing electrochemical platform.

Glycoconjugates found on cell walls of Candida species are fundamental for their pathogenicity. Laborious techniques have been employed to investigate the sugar composition of these microorganisms. Herein, we prepared a nanotool, based on the fluorescence of quantum dots (QDs) combined with the specificity of Cramoll lectin, to evaluate glucose/mannose profiles on three Candida species. The QDs-Cramoll conjugates presented specificity and bright fluorescence emission. The lectin preserved its biological activity after the conjugation process mediated by adsorption interactions. The labeling of Candida species was analyzed by fluorescence microscopy and quantified by flow cytometry. Morphological analyses of yeasts labeled with QDs-Cramoll conjugates indicated that C. glabrata (2.7 μm) was smaller when compared to C. albicans (4.0 μm) and C. parapsilosis sensu stricto (3.8 μm). Also, C. parapsilosis population was heterogeneous, presenting rod-shaped blastoconidia. More than 90% of cells of the three species were labeled by conjugates. Inhibition and saturation assays indicated that C. parapsilosis had a higher content of exposed glucose/mannose than the other two species. Therefore, QDs-Cramoll conjugates demonstrated to be effective fluorescent nanoprobes for evaluation of glucose/mannose constitution on the cell walls of fungal species frequently involved in candidiasis.

New nanoparticles containing biomaterials are emerging as versatile active platforms in a great number of applications, for example, as skin substitutes and therapeutic media. The present study describes the preparation of silver nanoparticles (AgNPs) embedded in agarose films and the impact of calcium ascorbate in the formation of ANPs as well as in the final properties of the films. Colloidal AgNPs were synthetized by two chemical reduction routes: (i) applying calcium ascorbate and NaBH4 and (ii) applying only NaBH4. AgNPs synthetized using NaBH4 showed sizes ranging from 5 to 18 nm while AgNPs were calcium ascorbate was used showed micrometer from 164 to 955 nm size. Films were prepared in three formulations: agarose control film (A1); agarose + AgNPs without calcium ascorbate (A2) and agarose + AgNPs with calcium ascorbate (A3). The characterization of films by SEM and EDS showed agarose agglomerates in A2 and unreacted calcium ascorbate crystals on surface of A3. Thus, the presence of calcium ascorbate influenced the properties of A3 film. In addition, the antimicrobial analysis showed a silver particles release dependence on the film composition and only the A3 presented activity against Staphylococcus aureus. The results found in this study open an important way for development of new biomaterials, economically competitive, and with medical application.

Leishmaniasis is a neglected disease present in more than 88 countries. The currently adopted chemotherapy faces challenges related to side effects and the development of resistance. Photodynamic therapy (PDT) is emerging as a therapeutic modality for cutaneous leishmaniasis. Zn(II) meso-tetrakis(N-ethylpyridinium-2-yl)porphyrin (ZnTE-2-PyP4+, ZnP) is a cationic, water-soluble, zinc porphyrin-based photosensitizer whose photodynamic effect on Leishmania braziliensis was analyzed by evaluating the number of visibly undamaged and motile cells, cell membrane integrity, mitochondrial membrane potential, and ultrastructural damage. Treatment of parasites with ZnP and light induced damage in up to 90% of L. braziliensis promastigote cells. Propidium iodide labeling suggested the loss of plasma membrane integrity. In samples treated with ZnP and light, a hyperpolarization of the mitochondrial membrane potential was also observed. Ultrastructural evaluation of promastigotes after photodynamic treatment indicated a loss of cytoplasmic material and the presence of vacuoles. Scanning electron microscopy showed wrinkling of the plasma membrane and a reduced cell volume. Additionally, the number of amastigotes per macrophage was reduced by about 40% after photodynamic application. The treatment showed no considerable toxicity against mammalian cells. Therefore, the results indicated that PDT associated with ZnTE-2-PyP4+ represents a promising alternative to cutaneous leishmaniasis treatment.

A fast and simple method for determination of six folate forms was developed through a study of standards and tested in New Zealand spinach (Tetragonia expansa) samples. A systematic evaluation of the standard solubilization conditions, pH, and molar concentration of the aqueous solvent of the mobile phase was performed and evaluated by means of chromatographic parameters: retention time (t r), capacity factor (k), alpha (α), and resolution (Rs). The optimal separation was achieved using phosphate buffer pH 2.0, 100 mmol L−1 and methanol in the ratio 85:15 isocratic mode using C18 Spherisorb ODS-2, 2.0 × 100 mm, 5 μm column, and DAD detection 200–400 nm. The following k values were obtained: 0.58, 1.17, 1.06, 1.98, 3.03, and 6.05, respectively, to tetrahydrofolate, 5-methyltetrahydrofolate, 10-formyltetrahydrofolate, 5-formyltetrahydrofolate, folic, and pteroic acid and α values of 1.81, 1.10, 1.69, 1.53, and 1.99. The elution order of the six folate forms is explained using the Lewis theory of acids and bases. This method was applied to samples of New Zealand spinach and monitored through chromatographic and spectral parameters obtained in the region of maximum absorption of λ. Moreover, the similarity and the purity of the peaks in standards and samples were investigated. For all folate forms, α and Rs values were above 1, except tetrahydrofolate. The similarity index was above 75 % and a purity above 90 % in samples, except for 10-formytetrahydrofolate, indicating that the method has good selectivity for complex matrices.

Camila G Andrade,1 Paulo E Cabral Filho,2 Denise PL Tenório,3 Beate S Santos,4 Eduardo IC Beltrão,1 Adriana Fontes,2 Luiz B Carvalho Jr1 1Keizo Asami Immunopathology Laboratory, 2Biophysics and Radiobiology Department, 3Fundamental Chemistry Department, 4Pharmaceutical Science Department, Federal University of Pernambuco, Recife, Pernambuco, Brazil Abstract: Cell surface glycoconjugates play an important role in differentiation/dedifferentiation processes and lectins are employed to evaluate them by several methodologies. Fluorescent probes are considered a valuable tool because of their ability to provide a particular view, and are more detailed and sensitive in terms of cell structure and molecular content. The aim of this study was to evaluate and compare the expression and distribution of glycoconjugates in normal human breast tissue, and benign (fibroadenoma), and malignantly transformed (invasive ductal carcinoma) breast tissues. For this, we used mercaptosuccinic acid-coated Cadmium Telluride (CdTe) quantum dots (QDs) conjugated with concanavalin A (Con A) or Ulex europaeus agglutinin I (UEA I) lectins to detect α-D-glucose/mannose and L-fucose residues, respectively. The QD-lectin conjugates were evaluated by hemagglutination activity tests and carbohydrate inhibition assays, and were found to remain functional, keeping their fluorescent properties and carbohydrate recognition ability. Fluorescence images showed that different regions of breast tissue expressed particular types of carbohydrates. While the stroma was preferentially and intensely stained by QD-Con A, ductal cells were preferentially labeled by QD-UEA I. These results indicate that QD-lectin conjugates can be used as molecular probes and can help to elucidate the glycoconjugate profile in biological processes. Keywords: nanoparticles, concanavalin A, Ulex europaeus, carbohydrates, mammary tissues

The application of fluorescent II-VI semiconductor quantum dots (QDs) as active photosensitizers in photodymanic inactivation (PDI) is still being evaluated. In the present study, we prepared 3 nm size CdTe QDs coated with mercaptosuccinic acid and conjugated them electrostatically with Zn(II) meso-tetrakis (N-ethyl-2-pyridinium-2-yl) porphyrin (ZnTE-2-PyP or ZnP), thus producing QDs-ZnP conjugates. We evaluated the capability of the systems, bare QDs and conjugates, to produce reactive oxygen species (ROS) and applied them in photodynamic inactivation in cultures of Candida albicans by irradiating the QDs and testing the hypothesis of a possible combined contribution of the PDI action. Tests of in vitro cytotoxicity and phototoxicity in fibroblasts were also performed in the presence and absence of light irradiation. The overall results showed an efficient ROS production for all tested systems and a low cytotoxicity (cell viability &gt, 90%) in the absence of radiation. Fibroblasts incubated with the QDs-ZnP and subjected to irradiation showed a higher cytotoxicity (cell viability &lt, 90%) depending on QD concentration compared to the bare groups. The PDI effects of bare CdTe QD on Candida albicans demonstrated a lower reduction of the cell viability (~1 log10) compared to bare ZnP which showed a high microbicidal activity (~3 log10) when photoactivated. The QD-ZnP conjugates also showed reduced photodynamic activity against C. albicans compared to bare ZnP and we suggest that the conjugation with QDs prevents the transmembrane cellular uptake of the ZnP molecules, reducing their photoactivity.

It is well known that the quantum confinement effects are closely related to the existence of different behavior for the same material composition. Due to the reduced size scale of the nanoparticles, the most part of their forming atoms are at the particle surface, which needs to be as stable as possible, to avoid phenomena such as dissolution and photodegradation. This way, methodologies for semiconductor nanoparticles obtention shall take into account the size, shape and energy of the final product. However, the relationship between these parameters is not yet clearly understood for nanometric systems, specially for those ones which are smaller than 20 nm. In this work, we present and discuss experimental and theoretical data obtained for nanoparticles of the semiconductor cadmium sulfide (CdS), in order to contribute for the understanding of the correlation between energetical and structural properties of nanometric systems in the quantum confinement regime.

We present and discuss the application of colloidal semiconductor quantum dots for diagnostic purposes, with special emphasis for cancer. We prepared and applied core–shell cadmium sulfide–cadmium hydroxide (CdS/Cd(OH)2) semiconductor quantum dots in aqueous medium. Tissue and cells labeling was evaluated by laser scanning confocal microscopy as well as by conventional fluorescence microscopy. The procedure presented in this work, shown to be a promising tool for fast, low-cost and precise cancer diagnostic protocols.

Semiconductor nanoparticles in the quantum confinement regime used as biolabels present many advantages over the other chemical species used as fluorophores. They are composed of 2000–6000 atoms rendering a far greater photostability and allowing for long time bioimaging experiments. In this work we present a synthetic route for the obtention of large quantities of highly fluorescent CdSe and CdTe/CdS core–shell nanocrystals based on aqueous colloidal chemistry. The methodologies were optimized and the systems were characterized by optical spectroscopy, transmission electronic microscopy and X-Ray diffractometry. The fluorescent biolabels were tested in live macrophages.

Semiconductor quantum dots based on II–VI materials may be prepared to develop good biolabeling properties. In this study we present some well-succeeded results related to the preparation, functionalization and bioconjugation of CdY (Y = S, Se and Te) to biological systems (live cells and fixed tissues). These nanostructured materials were prepared using colloidal synthesis in aqueous media resulting nanoparticles with very good optical properties and an excellent resistance to photodegradation.

Semiconductor colloidal quantum dots have been, for the past two decades, incorporated in a wide range of applications from catalysis and optical sensors to biolabels. For this reason, simple, cheap and reproducible routes of synthesis are the main goal of many research groups around the world. They seek the production of a very stable and extremely quantum efficient nanocrystal that can afford rough changes in the external environment. Silica capping is becoming a very common tool in the quest for a stable quantum dot, because of its strong and stable structure, this material provides a great insulator to the nanocrystal from the outside. The nanocrystal surface is not chemically favorable to the deposition of the bare silica shell, what demands a bifunctional molecule that provides the linkage between the core and the shell. In this work we present a comparison between several silanization methods of thiol capped CdSe and CdTe quantum dots, showing some simplifications of the routes and an application of the quantum dots produced as fluorescent cell markers in acquisition of confocal microscopy images.

In this study we use CdS/Cd(OH) 2 quantum dots functionalized with concanavalin-A (Con-A) lectin, specific to glucose/mannose residues, to investigate cell alterations regarding carbohydrate profile in human mammary tissues diagnosed as fibroadenoma (benign tumor). These particles were functionalized with glutaraldehyde and Con-A and incubated with tissue sections of normal and to Fibroadenoma, a benign type of mammary tumor. The tissue sections were deparafinized, hydrated in graded alcohol and treated with a solution of Evans Blue in order to avoid autofluorescence. The fluorescence intensity of QD-Con-A stained tissues showed different patterns, which reflect the carbohydrate expression of glucose/mannose in fibroadenoma when compared to the detection of the normal carbohydrate expression. The pattern of unspecific labeling of the tissues with glutaraldehyde functionalized CdS/Cd(OH) 2 quantum dots is compared to the targeting driven by the Con-A lectin. The preliminary findings reported here support the use of CdS/Cd(OH) 2 quantum dots as specific probes of cellular alterations and their use in diagnostics.

In this work we applied core-shell CdS/Cd(OH)2 quantum dots (QDs) as fluorescent labels in the Leishmania amazonensis protozoarium. The nanocrystals (8-9 nm) are obtained via colloidal synthesis in aqueous medium, with final pH=7 using sodium polyphosphate as the stabilizing agent. The surface of the particles is passivated with a cadmium hydroxide shell and the particle surface is functionalized with glutaraldehyde. The functionalized and non-functionalized particles were conjugated to Leishmania organisms in the promastigote form. The marked live organisms were visualized using confocal microscopy. The systems exhibit a differentiation of the emission color for the functionalized and non-functionalized particles suggesting different chemical interactions with the promastigote moieties. Two photon emision spectra (λ exc =795nm) were obtained for the promastigotes labeled with the functionalized QDs showing a significant spectral change compared to the original QDs suspension. These spectral changes are discussed in terms of the possible energy deactivation processes.

We present a simple and efficient method for marking living human red blood cells using CdS (Cadmium Sulfide) quantum dots (QDs). The nanocrystals were obtained via colloidal synthesis in aqueous medium with final pH=7 using sodium polyphosphate as the stabilizing agent. The methodology implementation is simple, do not requires additional capping layers nor narrow size QDs distribution. The synthesized nanoparticles were conjugated to monoclonal A anti-body. The resulting conjugates QDs/anti-A were incubated with human erythrocytes of blood groups A and O for 30 min at 37°C. The living cells in contact with the quantum dots maintained their properties for several days showing the low level of citotoxicity of the quantum dots. The conjugation of CdS QDs/anti-A show simultaneous red and green fluorescence when excited with 543 and 488 nm respectively. The efficiency of the conjugation QDs/anti-body to the erythrocytes, for each system, was monitored by confocal microscopy. The comparative analysis of the micrographs was done with the luminescence intensity maps of the samples obtained under constant capture conditions, such as, pinhole, filters, beam splitters and photomultiplier gain. The conjugates QDs/anti-A intensely marked group A erythrocytes and did not show any luminescence for group O erythrocytes, showing the sensitivity of the labeling procedure. In conclusion, we show the viability of the use of high luminescent and stable quantum dots as fluorescent labels for human erythrocytes with a methodology of simple implementation and the possibility to use them to distinguish different blood groups.

An electrochemical route for the generation of Te2− ions in an aqueous solution, with high yield and great stability, was developed and applied for the preparation of CdTe quantum dots (QDs) stabilized by negatively or positively charged ligands. An aqueous Cd(ClO4)2 solution containing mercaptopropionic acid (MPA) or cysteamine (Cys) stabilizers was directly added to the electrochemically generated aqueous telluride ion solution. This one pot process is quite reproducible and clean (no chemical reducing agents), allowing control of reduced species generation. The CdTe QDs showed good stability and high luminescence with particle average sizes of d = 3.1 nm (MPA) or 2.7 nm (Cys). Macrophages were efficiently labeled with MPA-capped CdTe QDs as well as with Cys-capped CdTe QDs.

We report a new methodology for red blood cell antigen expression determination by a simple labeling procedure employing luminescent semiconductor quantum dots. Highly luminescent and stable core shell cadmium sulfide/cadmium hydroxide colloidal particles are obtained, with a predominant size of 9 nm. The core-shell quantum dots are functionalized with glutaraldehyde and conjugated to a monoclonal anti-A antibody to target antigen-A in red blood cell membranes. Erythrocyte samples of blood groups A+, A2+, and O+ are used for this purpose. Confocal microscopy images show that after 30 min of conjugation time, type A+ and A2+ erythrocytes present bright emission, whereas the O+ group cells show no emission. Fluorescence intensity maps show different antigen expressions for the distinct erythrocyte types. The results obtained strongly suggest that this simple labeling procedure may be employed as an efficient tool to investigate quantitatively the distribution and expression of antigens in red blood cell membranes.

Semiconductor quantum dots (QDs) are nanometer-sized crystals with unique photochemical and photophysical properties that are not available from either isolated molecules or bulk solids. In comparison with organic dyes and fluorescent proteins, QDs are emerging as a new class of fluorescent labels with improved brightness, resistance against photobleaching, and multicolor fluorescence emission. These properties could improve the sensitivity of biological detection and imaging by at least 10 to 100 fold. Recent research has shown that QDs can be covalently linked with biorecognition molecules such as peptides, antibodies, nucleic acids and small molecules for use as fluorescent probes. The aim of the present study was to evaluate the application of a Cadmium Sulfide QDs method for the analysis of the expression and distribution of A antigen on living red blood cell surface. Highly luminescent and stable cadmium sulfide colloidal particles were obtained in the nanometer size range. Their surface was functionalized with commercially available monoclonal anti-A antibody via an one-pot glutaraldehyde cross linking procedure, followed by their conjugation to red cells. Samples were obtained from blood bank red cell units, of groups A (n=3), A2 (n=2) and O (n=3). Confocal microscopy (LSM 510 - Carl Zeiss, Jena, Germany) was used and for each cell type, the images were reproduced at least three times. The acquisition confocal image parameters, such as pinhole, filters, beam splitters and photomultiplier gain, were maintained constant to permit a comparative analysis of the luminescence intensity maps. Analyzing the intensity map of group A erythrocytes, a homogeneous emission profile was observed throughout the cell surface, suggesting a homogeneous distribution of A antigen. In contrast, the intensity map analysis of the confocal images showed a lower intensity pattern of the A2 group erythrocytes studied, suggesting different antigen expression on cell surface. The O group cells showed no emission. This result suggests that this new efficient, fast and simple labeling procedure may be employed as a quantitative tool to investigate the expression and distribution of molecules in red blood cells.

Liposomes are being studied as drug delivery systems, membrane models, as well as encapsulating systems of nanostructured materials for cellular imaging and diagnostics applications. Taking advantage of the use of Quantum Dots (QDs) as fluorescent probes over many conventional fluorescent dyes, several research groups are developing methods to encapsulate lypophilic and hydrophilic QDs into liposomes either in the aqueous compartment or in their bilayer in order to have a multifunctional system and provide therapy and diagnostics at the same time [1].In this work, Cadmium Telluride/Cadmium Sulfide (CdTe/CdS) quantum dots functionalized with mercaptopropionic acid (MPA) was synthesized in aqueous medium and characterized by optical spectroscopy and X-Ray diffractometry. Synthesis was carried out by adding Te−2 to a Cd+2/MPA refluxed for 7 h at ∼ 95°C. Cationic multifluorescent quantum dot liposomes (QD-Ls) formed by phosphatidylcholine (PC) and 1,2-dioleoyl-3-trimethylammonium-propane(DOTAP) (80:20) were prepared by lipid film hydration with pure water. QDs were encapsulated by a new method using freeze-and-thaw cycle. Liposomes (100 or 800 nm in diameter) encapsulating QDs were characterized by zeta potential, size distribution, fluorescence and transmission electron microscopy and show great potential for diagnostic purposes.[1] G. D. Bothun, A. E. Rabideau, and M. A. Stoner, J. Phys. Chem B Lett 2009 113, 7725-7728.Support: FACEPE, CNPq, INCT de Fotonica.

Fluorescent semiconductor nanocrystals called “quantum dots” (QDs) have been synthesized for applications in many areas, such as biomedical sciences as biosensors, disease diagnostics and cell labeling, as well as in electronics and optics. In this work, cadmium telluride (CdTe) QDs functionalized with mercaptosuccinic acid (MSA) was synthesized in aqueous medium by adding a solution of Te2- ions to a solution of cadmium perchlorate and MSA, followed by reflux under argon for 10 h at 95°C. These QDs were characterized by optical spectroscopy, X-ray diffractommetry and transmission electron microscopy (TEM). Cationic liposomes composed of egg phosphatidylcholine (PC) and 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP) were prepared by hydrating a lipid film at a lipid ratio of 80:20 respectively. This method consists in drying the lipids and the formation of a film under N2 for 30 min and drying it for 3 h under vacuum to remove residual solvent from the system. The hydration step was performed by adding pure water followed by stirring. The QDs have been synthesized previously encapsulated within these vesicles by the method of freezing and thawing. These vesicles were characterized by Zeta potential, Fluorescence Microscopy and TEM. Primary astrocytes from newborn Wistar rats (Pn 0 - 3, Pn = day of postnatal) were isolated and cultured in Dulbecco's Modified Eagle's Medium supplemented with F −12 (DMEM/F-12) and 10% fetal bovine serum. Upon 80% confluence, the culture medium was removed and fresh serum-free medium was added. Liposomes with and without QDs were tested in these cells with a high unspecific labeling character. The results demonstrate that liposomes present an effective entrapment of these fluorescent probes and are potential candidates for the development of diagnostics of pathologies related to glial tumor cells such as gliomas and glioblastomas.