Engineering of layer-by-layer coated capsules with the prospect of materials for efficient and directed electron transfer

Intermolecular electron transfer is investigated in a dye-doped polyelectrolyte (PE) multilayer film. Hollow PE capsules, with a mean diameter of 2 microm, were prepared by stepwise adsorption of a pyrene (PY)-labeled polyanion and various polycations onto charged colloids and subsequent dissolution of the colloidal core. The high concentration of dye molecules within the capsule wall and the control of the medium polarity on a nanometer length scale are proposed to facilitate light-induced charge separation over distances of a few nanometers. In particular, a PY-labeled poly(styrene sulfonate) (PSS-PY) has been synthesized and used as polyanion for the polyelectrolyte capsule preparation. A polarity gradient across the wall of the PE shells is assumed to be achieved by adsorbing diverse polycations at different film positions. The high effective film area followed by high optical density of the PE capsule solution enables time-resolved optical spectroscopy. Using pulsed excited state absorption (ESA) the transient absorption peaks of the radical anion and cation state of pyrene were measured, respectively. In the presence of additional electron donor (or acceptor) molecules in the capsule solution the pyrene anion (cation) is observed in the ESA spectra, while both transient states are seen if no additional molecules are present. These results are interpreted as an electron transfer from pyrene to the donor (acceptor) molecule or between two pyrene molecules. An asymmetry of the electron donor and electron acceptor efficiency was observed when multilayer shells were used that are supposed to carry an internal polarity gradient.     

Unveiling the chemistry behind bromination of quercetin: the ‘violet chromogen’

Bromination of quercetin with N-bromosuccinimide in neutral aqueous methanol occurs surprisingly in the electron-deficient A-ring only. Deprotonation of the acidic 7-OH is a major driver of this regioselective reaction. The increase of electron density makes in fact the quercetin anion suitable for an electrophilic attack by bromine at positions 8 and 6. Several pieces of evidence (NMR spectra and H/D exchange) are presented to substantiate the mechanism advanced. Bromoquinones/quinomethides produced in excess of N-bromosuccinimide are responsible for the formation of a stable ‘violet chromogen’.     

Stereoselective synthesis of constrained norbornane-derived spiro-β-lactams

New enantiopure polycyclic norbornane-derived spiro-β-lactams were synthesized by means of a Staudinger ketene–imine reaction between unsymmetrical bicyclic chiral ketenes, generated from differently substituted norbornane carboxylic acids, and (E)-N-benzyl-N-(phenylmethylene)amine, with high yields and moderate to good stereoselectivities. The diastereoisomeric results were rationalized taking into account the increasing steric encumbrance present on the norbornane skeleton and the stability of the products. The configurations of the newly formed stereocenters of spiro-β-lactams were assigned on the basis of 2D NMR experiments and X-ray analysis. Spiro-β-lactams were subjected to acid hydrolysis obtaining the corresponding norbornane-derived β-amino acids.     

A New Alkaliphilic Cold-Active Esterase from the Psychrophilic Marine Bacterium Rhodococcus sp.: Functional and Structural Studies and Biotechnological Potential

The special features of cold-adapted lipolytic biocatalysts have made their use possible in several industrial applications. In fact, cold-active enzymes are known to be able to catalyze reactions at low temperatures, avoiding side reactions taking place at higher temperatures and preserving the integrity of products. A lipolytic gene was isolated from the Arctic marine bacterium Rhodococcus sp. AW25M09 and expressed in Escherichia coli as inclusion bodies. The recombinant enzyme (hereafter called RhLip) showed interesting cold-active esterase activity. The refolded purified enzyme displayed optimal activity at 30 °C and was cold-active with retention of 50 % activity at 10 °C. It is worth noting that the optimal pH was 11, and the low relative activity below pH 10 revealed that RhLip was an alkaliphilic esterase. The enzyme was active toward short-chain p-nitrophenyl esters (C2–C6), displaying optimal activity with the butyrate (C4) ester. In addition, the enzyme revealed a good organic solvent and salt tolerance. These features make this an interesting enzyme for exploitation in some industrial applications.     

Simultaneous determination of five common additives in insulating mineral oils by high‐performance liquid chromatography with ultraviolet and coulometric detection

Dielectric mineral oils are used to impregnate power transformers and large electrical apparatus, acting as both liquid insulation and heat dissipation media. Antioxidants and passivators are frequently added to mineral oils to enhance oxidation stability and reduce the electrostatic charging tendency, respectively. Since existing standard test methods only allow analysis of individual additives, new approaches are needed for the detection of mixtures. For the first time we investigate and discuss the performance of analytical methods, which require or do not require extraction as sample pretreatment, for the simultaneous reversed‐phase high‐performance liquid chromatography determination of passivators (benzotriazole, Irgamet^® 39) and antioxidants (N‐phenyl‐1‐naphtylamine, 2,6‐di‐tert‐butylphenol, 2,6‐di‐tert‐butyl‐p‐cresol), chosen for their presence in marketed oils. Quick easy, cheap, effective, rugged and safe and solid phase extractions were evaluated as sample pretreatments. Direct sample‐injection was also studied. Ultraviolet spectrophotometry and direct‐current coulometry detection were explored. As less prone to additive concentrations variability, the direct‐injection high‐performance liquid chromatography with ultraviolet and coulometric detection method was validated through comparison with Standard Method IEC 60666 and through an ASTM interlaboratory proficiency test. Obtained detection limits are (mg kg^?1): benzotriazole (2.8), Irgamet^® 39 (13.8), N‐phenyl‐1‐naphtylamine (11.9), 2,6‐di‐tert‐butylphenol (13.1), 2,6‐di‐tert‐butyl‐p‐cresol (10.2). Simultaneous determination of selected additives was possible both in unused and used oils, with good precision and accuracy.     

Characterization of a cold-active and salt tolerant esterase identified by functional screening of Arctic metagenomic libraries

Background

The use of metagenomics in enzyme discovery constitutes a powerful approach to access to genomes of unculturable community of microorganisms and isolate novel valuable biocatalysts for use in a wide range of biotechnological and pharmaceutical fields.

Results

Here we present a novel esterase gene (lip3) identified by functional screening of three fosmid metagenomic libraries, constructed from three marine sediment samples. The sequenced positive fosmid revealed an enzyme of 281 amino acids with similarity to class 3 lipases. The 3D modeling of Lip3 was generated by homology modeling on the basis of four lipases templates [PDB ID: 3O0D, 3NGM, 3G7N, 2QUB] to unravel structural features of this novel enzyme. The catalytic triad of Lip3 was predicted to be Asp207, His267 and the catalytic nucleophile Ser150 in a conserved pentapeptide (GXSXG). The 3D model highlighted the presence of a one-helix lid able to regulate the access of the substrate to the active site when the enzyme binds a hydrophobic interface. Moreover an analysis of the external surface of Lip3 model showed that the majority of the surface regions were hydrophobic (59.6 %) compared with homologous lipases (around 35 %) used as templates. The recombinant Lip3 esterase, expressed and purified from Escherichia coli, preferentially hydrolyzed short and medium length p-nitrophenyl esters with the best substrate being p-nitrophenyl acetate. Further characterization revealed a temperature optimum of 35 °C and a pH optimum of 8.0. Lip3 exhibits a broad temperature stability range and tolerates the presence of DTT, EDTA, PMSF, β-mercaptoethanol and high concentrations of salt. The enzyme was also highly activated by NaCl.

Conclusions

The biochemical characterization and homology model reveals a novel esterase originating from the marine Arctic metagenomics libraries with features of a cold-active, relatively thermostable and highly halotolerant enzyme. Taken together, these results suggest that this esterase could be a highly valuable candidate for biotechnological applications such as organic synthesis reactions and cheese ripening processes.

     

Dioxouranium(VI)-carboxylate complexes. Speciation of UO2(2+)-1,2,3-propanetricarboxylate system in NaCl(aq) at different ionic strengths and at t=25 degrees C

The formation constants of dioxouranium(VI)-1,2,3-propanetricarboxylate [tricarballylate (3-), TCA] complexes were determined in NaCl aqueous solutions at 0 < or = I/mol L(-1) < or = 1.0 and t=25 degrees C, by potentiometry, ISE-[H+] glass electrode. The speciation model obtained at each ionic strength includes the following species: ML-, MLH0, ML2(4-) and ML2H3- (M = UO2(2+) and L = TCA). The dependence on ionic strength of protonation constants of 1,2,3-propanetricarboxylate and of the metal-ligand complexes was modeled by the SIT (Specific ion Interaction Theory) approach and by the Pitzer equations. The formation constants at infinite dilution are [for the generic equilibrium p UO22+ + q (L3-) + r H+ = (UO2(2+))p(L)qHr(2p-3q+r); betapqr]: log beta110 = 6.222 +/- 0.030, log beta111 = 11.251 +/- 0.009, log beta121 = 7.75 +/- 0.02, log beta121 = 14.33 +/- 0.06. The sequestering ability of 1,2,3-propanetricarboxylate towards UO2(2+) was quantified by using a sigmoid Boltzman type equation.     

New Oxaliplatin-Pyrophosphato Analogs with Improved In Vitro Cytotoxicity

Two new Pt(II)-pyrophosphato complexes containing the carrier ligands cis-1,3-diaminocyclohexane (cis-1,3-DACH) and trans-1,2-diamine-4-cyclohexene (1,2-DACHEX), variants of the 1R,2R-diaminocyclohexane ligand present in the clinically used Pt-drug oxaliplatin, have been synthesized with the aim of developing new potential antitumor drugs with high bone tropism. The complexes are more stable at physiological pH than in acid conditions, with Na₂[Pt(pyrophosphato)(cis-1,3-DACH)] (1) slightly more stable than [Pt(dihydrogenpyrophosphato)(1,2-DACHEX)] (2). The greater reactivity at acidic pH ensures a greater efficacy at the tumor site. Preliminary NMR studies indicate that 1 and 2 react slowly with 5’-GMP (used as a model of nucleic acids), releasing the pyrophosphate ligand and affording the bis 5’-GMP adduct. In vitro cytotoxicity assays performed against a panel of four human cancer cell lines have shown that both compounds are more active than oxaliplatin. Flow cytometry studies on HCT116 cells showed that the pyrophosphato compounds with the non-classical 1,3- and 1,4-diaminocyclohexane ligands (1 and 4) are the most capable to induce cells’ death by apoptosis and necrosis.     

Diphenylalanine Motif Drives Self-Assembling in Hybrid PNA-Peptide Conjugates

Peptides and nucleic acids can self-assemble to give supramolecular structures that find application in different fields, ranging from the delivery of drugs to the obtainment of materials endowed with optical properties. Forces that stabilize the “suprastructures” typically are hydrogen bonds or aromatic interactions; in case of nucleic acids, Watson-Crick pairing drives self-assembly while, in case of peptides, backbone hydrogen bonds and interactions between aromatic side chains trigger the formation of structures, such as nanotubes or ribbons. Molecules containing both aromatic peptides and nucleic acids could in principle exploit different forces to self-assemble. In this work we meant to investigate the self-assembly of mixed systems, with the aim to understand which forces play a major role and determine formation/structure of aggregates. We therefore synthesized conjugates of the peptide FF to the peptide nucleic acid dimer “gc” and characterized their aggregates by different spectroscopic techniques, including NMR, CD and fluorescence.     

Retention of heavy metal ions on SBA-15 mesoporous silica functionalised with carboxylic groups

Three samples of SBA-15 functionalised with -(CH(2))(3)COOH groups have been prepared by co-condensation, starting from solutions of TEOS and 4-(triethoxysilyl)butyronitrile, acting as -(CH(2))(3)COOH precursor, of different molar compositions. Materials were characterised by X-ray diffraction, nitrogen adsorption, and FT-IR spectroscopy. The pK(a) and the acidic capacity were measured for all samples by potentiometric titration. The acidic capacity increases with increasing amount of -COOH precursor in the synthesis mixture only up to 10% molar of total alkoxysilane. The value for the pK(a)(4.75) is independent of the acidic capacity of the material. The sample prepared starting from an amount of -COOH precursor equal to 10% molar of total alkoxysilane was chosen to test selective interactions with heavy metals of environmental importance (Ag(+), Cd(2+), Fe(3+), Cu(2+), Zn(2+)) at different pH values and ionic strengths. The significant and selective adsorption exhibited by the material has been exploited in a preliminary cation-exchange chromatographic application showing the possibility of eluting the metal ions at different retention times.