A facile method to synthesize mussel-inspired polydopamine nanospheres as an active template for in situ formation of biomimetic hydroxyapatite.

In this study, Mussel-inspired polydopamine (PDA) nanospheres were synthesized via spontaneous oxidative polymerization of dopamine hydrochloride (dopa-HCl) in a deionized water-alcohol mixed solvent at room temperature and atmospheric air, under alkaline condition. Field-emission scanning electron microscopy (FE-SEM) demonstrated production of sphere-like shape with a smooth surface and tunable size, while monodispersity increased by utilizing isopropanol instead of ethanol owing to lower Ra values based on Hansen solubility parameter (HSP) theory. Dropwise addition of monomer played an undeniable role in the fabrication of uniform and smaller spheres. The difference of the charge repulsion of constructs in the range of pH led to different dispersive behavior in a variety of solvents, exhibiting versatile applications. The presence of active functional groups on the surface of PDA spheres made them an appropriate option for PDA-assisted biomimetic mineralization of hydroxyapatite (HA), which is the result of the interaction between abundant catecholamine moieties in PDA and Ca+2 ions in simulated body fluid. Bio-adhesive nature of PDA in water and the presence of amino and hydroxyl functional groups support desirable L929 mouse fibroblast cell spreading. The viability of >90% fibroblast cells proved the biocompatibility of polymerized structure. All the achievements indicated that PDA nanospheres provide a biocompatible and bioactive template for green synthesizing hydroxyapatite and the innovative basis for further tissue engineering applications.

Ionic asymmetry and solvent excluded volume effects on spherical electric double layers: a density functional approach.

In this article, we present a classical density functional theory for electrical double layers of spherical macroions that extends the capabilities of conventional approaches by accounting for electrostatic ion correlations, size asymmetry, and excluded volume effects. The approach is based on a recent approximation introduced by Hansen-Goos and Roth for the hard sphere excess free energy of inhomogeneous fluids [J. Chem. Phys. 124, 154506 (2006); Hansen-Goos and Roth, J. Phys.: Condens. Matter 18, 8413 (2006)]. It accounts for the proper and efficient description of the effects of ionic asymmetry and solvent excluded volume, especially at high ion concentrations and size asymmetry ratios including those observed in experimental studies. Additionally, we utilize a leading functional Taylor expansion approximation of the ion density profiles. In addition, we use the mean spherical approximation for multi-component charged hard sphere fluids to account for the electrostatic ion correlation effects. These approximations are implemented in our theoretical formulation into a suitable decomposition of the excess free energy which plays a key role in capturing the complex interplay between charge correlations and excluded volume effects. We perform Monte Carlo simulations in various scenarios to validate the proposed approach, obtaining a good compromise between accuracy and computational cost. We use the proposed computational approach to study the effects of ion size, ion size asymmetry, and solvent excluded volume on the ion profiles, integrated charge, mean electrostatic potential, and ionic coordination number around spherical macroions in various electrolyte mixtures. Our results show that both solvent hard sphere diameter and density play a dominant role in the distribution of ions around spherical macroions, mainly for experimental water molarity and size values where the counterion distribution is characterized by a tight binding to the macroion, similar to that predicted by the Stern model.

Charge density and electrostatic potential analyses in paracetamol.

The electron density of monoclinic paracetamol was derived from high-resolution X-ray diffraction at 100 K. The Hansen-Coppens multipole model was used to refine the experimental electron density. The topologies of the electron density and the electrostatic potential were carefully analyzed. Numerical and analytical procedures were used to derive the charges integrated over the atomic basins. The highest charge magnitude (-1.2 e) was found for the N atom of the paracetamol molecule, which is in agreement with the observed nucleophilic attack occurring in the biological media. The electric field generated by the paracetamol molecule was used to calculate the atomic charges using the divergence theorem. This was simultaneously applied to estimate the total electrostatic force exerted on each atom of the molecule by using the Maxwell stress tensor. The interaction electrostatic energy of dimers of paracetamol in the crystal lattice was also estimated.

Development of an environmentally acceptable volatile organic compounds recovery and reuse system based on vacuum spray flash with dibasic acid esters as absorbents.

We used dibasic acid esters as the absorbent for a volatile organic compounds (VOC) recovery and reuse system geared mainly toward VOCs from painting and presswork. The system consists of three elements: absorption using dibasic acid esters, separation by vacuum spray flash method, and recovery by condensation. The concentration of both ethyl acetate (EtAc) and isopropyl alcohol (IPA) decreased around 90% at room temperature. Vacuum spray flash was found as a useful process for cost-effective and energy-effective VOC removal system. The difference in the dissolubility parameters (Hansen constant) between dibasic acid esters and VOCs and the boiling point of VOCs was the main contributor to the separation behavior.

Heterologous expression implicates a GATA factor in regulation of nitrogen metabolic genes and ion homeostasis in the halotolerant yeast Debaryomyces hansenii.

The yeast Debaryomyces hansenii has a remarkable capacity to proliferate in salty and alkaline environments such as seawater. A screen for D. hansenii genes able to confer increased tolerance to high pH when overexpressed in Saccharomyces cerevisiae yielded a single gene, named here DhGZF3, encoding a putative negative GATA transcription factor related to S. cerevisiae Dal80 and Gzf3. Overexpression of this gene in wild-type S. cerevisiae increased caffeine and rapamycin tolerance, blocked growth in low glucose concentrations and nonfermentable carbon sources, and resulted in lithium- and sodium-sensitive cells. Sensitivity to salt could be attributed to a reduced cation efflux, most likely because of a decrease in expression of the ENA1 Na(+)-ATPase gene. Overexpression of DhGZF3 did not affect cell growth in a gat1 mutant but was lethal in the absence of Gln3. These are positive factors that oppose both Gzf3 and Dal80. Genome-wide transcriptional profiling of wild-type cells overexpressing DhGZF3 shows decreased expression of a number of genes that are usually induced in poor nitrogen sources. In addition, the entire pathway leading to Lys biosynthesis was repressed, probably as a result of a decrease in the expression of the specific Lys14 transcription factor. In conclusion, our results demonstrate that DhGzf3 can play a role as a negative GATA transcription factor when expressed in S. cerevisiae and that it most probably represents the only member of this family in D. hansenii. These findings also point to the GATA transcription factors as relevant elements for alkaline-pH tolerance.

Identification of proteins induced by polycyclic aromatic hydrocarbon in Mycobacterium vanbaalenii PYR-1 using two-dimensional polyacrylamide gel electrophoresis and de novo sequencing methods.

Protein profiles of Mycobacterium vanbaalenii PYR-1 grown in the presence of high-molecular-weight polycyclic aromatic hydrocarbons (HMW PAHs) were examined by two-dimensional gel electrophoresis (2-DE). Cultures of M. vanbaalenii PYR-1 were incubated with pyrene, pyrene-4,5-quinone (PQ), phenanthrene, anthracene, and fluoranthene. Soluble cellular protein fractions were analyzed and compared, using immobilized pH gradient (IPG) strips. More than 1000 gel-separated proteins were detected using a 2-DE analysis program within the window of isoelectric point (pI) 4-7 and a molecular mass range of 10-100 kDa. We observed variations in the protein composition showing the upregulation of multiple proteins for the five PAH treatments compared with the uninduced control sample. By N-terminal sequencing or mass spectrometry, we further analyzed the proteins separated by 2-DE. Due to the lack of genome sequence information for this species, protein identification provided an analytical challenge. Several PAH-induced proteins were identified including a catalase-peroxidase, a putative monooxygenase, a dioxygenase small subunit, a small subunit of naphthalene-inducible dioxygenase, and aldehyde dehydrogenase. We also identified proteins related to carbohydrate metabolism (enolase, 6-phosphogluconate dehydrogenase, indole-3-glycerol phosphate synthase, and fumarase), DNA translation (probable elongation factor Tsf), heat shock proteins, and energy production (ATP synthase). Many proteins from M. vanbaalenii PYR-1 showed similarity with protein sequences from M. tuberculosis and M. leprae. Some proteins were detected uniquely upon exposure to a specific PAH whereas others were common to more than one PAH, which indicates that induction triggers not only specific responses but a common response in this strain.

Metal ion transport and regulation in Mycobacterium tuberculosis.

The regulation of metal ion concentrations is central to the physiology of the interaction between pathogenic bacteria and their hosts. Apart from the NRAMP orthologue, MntH, metal ion transporters in Mycobacterium tuberculosis have not been studied. Mn, the physiological substrate of MntH in other bacteria, may play an important role as a structural and redox-active cofactor in a wide range of metabolic processes. Fe, Cu and Zn play structural and catalytic roles in metalloenzymes involved in oxidative stress responses. Fe and Mg are required for growth in macrophages. Genomic analyses reveal 28 sequences encoding a broad repertoire of putative metal ion transporters (or transporter subunits), representing 24% of all transporters in this organism. These comprise 8 families of secondary active transporters and 3 families of primary active transporters, including 12,P, type ATPases. Potential metal ion specificities include K+, Na+, Cu2+, Cd2+, Zn2+, Mn2+, Mg2+, Ca2+, Co2+, Ni2+, Fe2+/3+, Hg2+, AsO2- and AsO4(2-). 17 of these transporters are also encoded as complete open reading frames in Mycobacterium leprae, suggesting a role in intracellular survival. Iron transcriptionally regulates a diverse set of genes via the iron-dependent DNA-binding proteins, Fur and IdeR. Changes in Fe and Mg concentrations signal entry into the intracellular compartment and potentially trigger up-regulation of virulence determinants. The plethora of putative transport systems encoded by the M. tuberculosis genome contrasts strikingly with the paucity of experimental data on these systems. The detailed analysis of the temporal pattern of M. tuberculosis transporter gene expression during infection will provide important insights into the basic biology of intracellular parasitism and may help to shape novel therapeutic strategies.

Spectrophotometric determination of dapsone in pharmaceutical products using sodium 1,2-naphthoquinone-4-sulfonic as the chromogenic reagent.

Spectrophotometric determination of dapsone is described. The dapsone reacts with sodium 1,2-naphthoquinone-4-sulfonic in pH 6.98 buffer solution to form a salmon pink compound, and its maximum absorption wavelength is at 525 nm, epsilon525=3.68 x 10(4) l mol(-1) cm(-1). The absorbance of dapsone from 0.40 to 10 microg ml(-1) obeys Beer's law. The linear regression equation of the calibration graph is C=0.2334 A + 0.01288, with a linear regression correlation coefficient of 0.9998, the detection limit is 0.24 microg ml(-1), and recovery is from 99.2 to 102.4%. Effects of pH, surfactant, organic solvents, foreign ions, and standing time on the determination of dapsone have been examined. This method is simple and can be used for the determination of dapsone in injection solution of dapsone. The results obtained by this method agreed with those by the official method (dead-stop titration method [The Chinese Pharmacopoeia, Pharmacopoeia Commission, Ministry of Health, vol. 2, fifth ed., PRC Chemical Industry Press, Beijing, 2000, p.720]).

Criteria to design green enzymatic processes in ionic liquid/supercritical carbon dioxide systems.

Five different ionic liquids (ILs) based on quaternary ammonium cations, with functional side chains ((3-hydroxypropyl)-trimethyl-, (3-cyanopropyl)-trimethyl-, butyl-trimethyl-, (5-cyanopentyl)-trimethyl- and hexyl-trimethyl-) associated with the same anion (bis(trifluoromethane)sulfonyl amide)), were synthesized, and their suitability for Candida antarctica lipase B (CALB)-catalyzed ester synthesis in IL/supercritical carbon dioxide (scCO(2)) biphasic systems was assayed. Catalytic efficiency of the system has been analyzed as a function of both enzyme properties and mass-transfer phenomena criteria. First, the suitability of these ILs as enzymic reaction media was tested for the kinetic resolution of rac-phenylethanol. All ILs were found to be suitable media for enzyme catalysis, the best catalytic parameter (5.3 U/mg specific activity, 94.9% selectivity) being obtained for the (5-cyanopentyl)-trimethylammonium. Second, enzyme stability in all of the ILs was studied at 50 degrees C over a period of 50 days, and data were analyzed by a two-step kinetic deactivation model. All of the ILs were shown to act as stabilizing agents with respect to hexane, producing an increase in the free energy of deactivation (to 25 kJ/mol protein) and an improvement in the half-life time of the enzyme (2000-fold), which agrees with the observed increased hydrophobicity of the cation alkyl side chain (measured by Hansen's solubility parameter, delta). By using two different CALB-IL systems with different hydrophobicity in the cation, continuous processes to synthesize six different short chain alkyl esters (butyl acetate, butyl propionate, butyl butyrate, hexyl propionate, hexyl butyrate, and octyl propionate) in scCO(2) at 10 MPa and 50 degrees C were carried out. Both rate-limiting parameters (synthetic activity and scCO(2)-ILs mass-transfer phenomena) were related with the delta-parameter of the ILs-alkyl chain and reagents.

Characterization of cell envelope-associated proteinases of thermophilic lactobacilli.

The proteolytic activities of two natural isolates of thermophilic lactobacilli, Lactobacillus acidophilus BGRA43 and Lact. delbrueckii BGPF1, and Lact. acidophilus CH2 (Chr. Hansen's strain) and Lact. acidophilus V74 (Visby's strain), were compared. Results revealed that optimal pH for all four proteinases is 6.5, whereas temperature optimum varied among proteinases. Determination of caseinolytic activity done under optimal conditions for each strain revealed that the CH2 and V74 proteinases completely hydrolysed both alphaS1-casein and beta-casein, showing very low activity towards kappa-casein. The BGPF1 proteinase completely hydrolysed only beta-casein. The BGRA43 proteinase completely hydrolysed all three casein fractions. The proteolytic activities of whole cells were inhibited by serine proteinase inhibitors, suggesting that all four strains produce serine proteinases. DNA-DNA hybridization and PCR analysis showed that BGPF1 contains the prtB-like proteinase gene. Characterized thermophilic strains BGPF1 and BGRA43 were successfully used as starter cultures for production of yoghurt and acidophilus milk, respectively.

Pigment producing yeasts involved in the brown surface discoloration of ewes' cheese.

The appearance of a brown surface discoloration on Portuguese ewes' cheese has never previously been reported on. The regular occurrence of this defect over the past few years has caused serious financial losses to producers, which has led to growing interest in its study. This paper describes a preliminary approach to the problem, based on the hypothesis that pigment producing yeasts are involved. From a group of 51 yeast strains isolated from a number of brown cheese rinds, it was possible to distinguish four pigment producing groups: group I (12 strains), produced an extracellular brown pigment from tyrosine and alkalised the tested media; group II (21 strains), produced a diffusible, reddish-brown pigment from resorcinol and alkalised the tested media; group III (three strains), alkalised the tested media without producing any pigments; group IV (15 strains), neither produced pigments nor alkalised the media. Yarrowia lipolytica and Candida catenulata type strains were also tested and their behaviour was similar to the strains in groups I and IV, respectively. The Filobasidiella neoformans type strain was distinct from all the other groups. The identification methods used for some strains in groups I, II and III suggest that Yarrowia lipolytica species is common to all strains in group I, and that Debaryomyces hansenii is present in both groups II and III. A study of the effect of several metal ions on the production of the brown pigment from tyrosine indicated Mn2+ to be a strong activator. Evidence is provided suggesting that the browning process may be related to tyrosine Yarrowia lipolytica metabolism.

Mechanisms of phagocytosis of Mycobacterium leprae and other mycobacteria by human oligodendroglial cells.

The mechanisms by which human oligodendroglial cells, KG-1-C cells, phagocytose mycobacteria, especially Mycobacterium leprae, were studied. The ability of glial cells to phagocytose M. leprae was inhibited by azide, dinitrophenol (inhibitors of oxidative phosphorylation), and iodoacetamide but not fluoride (both are inhibitors of glycolysis). Thus, the energy metabolism dependency is somewhat different from that of peritoneal macrophages and polymorphonuclear leukocytes, the phagocytic capacities of which are mainly dependent on glycolysis. Phagocytosis of M. leprae by KG-1-C cells was markedly suppressed by a microfilament inhibitor (cytochalasin B) but not microtubule inhibitors (colchicine and vinblastine), as with macrophages. The phagocytosis of M. leprae by KG-1-C cells was dependent on the lipid and somewhat on the sugar ligands of the organism. Moreover, the phagocytosis of a given mycobacterium by KG-1-C cells correlated well with its hydrophobicity, thus revealing the importance of some lipid moieties on the surface of bacteria in the establishment of rigid binding interaction of bacteria with KG-1-C cells, before the onset of engulfment. Electric charge of a given microorganism did not correlate with its phagocytosis by KG-1-C cells.

The most indolent ulcers of the skin treated with porcine xenografts and silver ions.

Cutaneous ulcers associated with Hansen's disease have long been recognized as among the most indolent of all ulcers. We have successfully treated such ulcers with porcine xenograft, achieving healing in ulcers which had been present for as much as three decades. These results suggest that this regimen may be applicable to other cutaneous ulcers wherever skin is missing. The newly available antibacterial effects of silver-impregnated porcine xenografts should have the result of increasing the effectiveness of the treatment still further.