Nanoscale organization in the fluorinated room temperature ionic liquid: Tetraethyl ammonium (trifluoromethanesulfonyl)(nonafluorobutylsulfonyl)imide.

Fluorinated Room Temperature Ionic Liquids (FRTILs) are a branch of ionic liquids that is the object of growing interest for a wide range of potential applications, due to the synergic combination of specifically ionic features and those properties that stem from fluorous tails. So far limited experimental work exists on the micro- and mesoscopic structural organization in this class of compounds. Such a work is however necessary to fully understand morphological details at atomistic level that would have strong implications in terms of bulk properties. Here we use the synergy between X-ray and neutron scattering together with molecular dynamics simulations to access structural details of a technologically relevant FRTIL that is characterised by an anion bearing a long enough fluorinated tail to develop specific morphological features. In particular, we find the first experimental evidence that in FRTILs bearing an asymmetric bis(perfluoroalkyl)sulfonyl-imide anion, fluorous side chains tend to be spatially segregated into nm-scale spatial heterogeneities. This feature together with the well-established micro-segregation of side alkyl chains in conventional RTILs leads to the concept of triphilic ILs, whose technological applications are yet to be fully developed.

Direct experimental observation of mesoscopic fluorous domains in fluorinated room temperature ionic liquids.

Fluorinated room temperature ionic liquids (FRTILs) represent a class of solvent media that are attracting great attention due to their IL-specific properties as well as features stemming from their fluorous nature. Medium-to-long fluorous tails constitute a well-defined apolar moiety in the otherwise polar environment. Similarly to the case of alkyl tails, such chains are expected to result in the formation of self-assembled fluorous domains. So far, however, no direct experimental observation has been made of the existence of such structural heterogeneities on the nm scale. We report here the first experimental evidence of the existence of mesoscopic spatial segregation of fluorinated domains, on the basis of highly complementary X-ray and neutron scattering data sets (highlighting the importance of the latter probe) and NMR spectroscopy. Data are interpreted using atomistic molecular dynamics simulations, emphasizing the existence of a self-assembly mechanism that delivers segregated fluorous domains, where preferential solubilisation of fluorinated compounds can occur, thus paving the way for several smart applications.

Analysis of IL-6, IL-10 and IL-17 genetic polymorphisms as risk factors for sepsis development in burned patients.

Infection risk, sepsis and mortality after severe burn are primarily determined by patient age, burn size, and depth. Whether genetic differences contribute to otherwise unexpected variability in outcomes is unknown. We sought to determine whether there was an association between IL-6, IL-10 and IL-17 polymorphisms with cytokine production and development of sepsis. We evaluated 71 patients with burns ≥15% TBSA and 109 healthy subjects. The genotypes of IL-6 (-174C/G), IL-10 (-819C/T and -1082A/G) and IL-17 (7488T/C) polymorphisms were identified applying polymerase chain reaction protocols. The cytokine levels in serum were determined with enzyme-linked immunoabsorbent assays. Our results demonstrated no significant differences in the genotype frequencies studied between burn patients and healthy subjects. No significant associations were found among IL-6 and IL-17F genotypes and the related cytokine serum levels. Only IL-10 promoter -1082GG genotype was related to an increased IL-10 production in burned patients. In addition, septic subjects bearing -1082G/G genotype have shown the highest and non-septic bearing -1082A/* genotypes the lowest IL-10 serum levels. All together these data seem to indicate that genetically determined individual difference in IL-10 production might influence the susceptibility to septic complications in burned patients and suggest that these markers might be useful in burned patient management.

Excess thermodynamic properties in mixtures of a representative room-temperature ionic liquid and acetonitrile.

The nature of the interactions between a representative room-temperature ionic liquid, namely 1-butyl-3-methyl imidazolium tetrafluoroborate ([BMIM][BF(4)]) and a common organic solvent, acetonitrile (CH(3)CN) has been investigated by means of Brillouin light scattering, over the whole concentration range and in the temperature range from -20 to 45 degrees C. Negative deviations from the ideal behavior of both molar volumes and adiabatic compressibility have been observed. This result has been interpreted within the framework of a well-established theoretical model, namely a nonadditive hard-sphere mixture. Despite that similar findings were rationalized in terms of enhanced interactions between molecules, a more detailed analysis of excess thermodynamic functions indicates that they are mainly due to excluded volume effects and that the differences in local intermolecular interactions act as higher order contributions: we have found that this can be a general feature of liquid mixtures. On this basis we present a reconsideration for excess thermodynamic data and for their role in providing direct information on intermolecular interactions.

Alanine blends for ESR measurements of thermal neutron fluence in a mixed radiation field.

In this paper, the results of a study on the electron spin resonance (ESR) dosimetry to measure thermal neutron fluence in a mixed radiation field (neutron and photons) are presented. The ESR responses of alanine dosemeters with different additives are compared. In particular, the (10)B-acid boric and the Gd-oxide were chosen to enhance the sensitivity of alanine dosemeters to thermal neutrons. Irradiations were carried out inside the thermal column of the TAPIRO reactor of the ENEA center, Casaccia Rome. The main results are a greater neutron sensitivity and a smaller lowest detectable fluence for the dosemeters with gadolinium than for dosemeters of alanine with (10)B, which is well known to be much more sensitive to thermal neutrons than simple alanine.

Study of the glow curves of TLD exposed to thermal neutrons.

The glow curves of thermoluminescent dosimeters (TLD600, TLD700 and MCP), exposed to a mixed field of thermal neutrons and gamma photons are analysed. The fluence values of thermal neutrons used, comparable with those used in radiotherapy, allow one to define the reliability of the TLDs, in particular the most sensitive MCP, in this radiation field and to get information on the dose absorbed values. The glow curves obtained have been deconvoluted using general order kinetics and the observed differences for the different LET components have been analysed. In particular, the ratio of the n(0) parameter of two different peaks seems to allow to discriminate the different contributions of neutrons and gamma photons in the beam.

Impaired reinnervation in HIV infection following experimental denervation.

OBJECTIVE: To determine whether abnormalities in peripheral nerve regeneration are present in HIV-infected individuals. DESIGN: We studied human epidermal nerve fiber reinnervation by collateral and regenerative sprouting using validated cutaneous nerve injury models in healthy control subjects and those infected with HIV. Characteristics associated with the degree of reinnervation were identified. METHODS: Seventy-one healthy volunteers and 22 HIV+ individuals underwent topical capsaicin treatment at the distal lateral thigh resulting in complete epidermal denervation. Regenerative sprouting was quantified in skin biopsies at intervals up to 100 days. Five healthy subjects and 5 HIV+ individuals underwent a 3-mm distal thigh punch skin biopsy to create an intracutaneous axotomy, followed 2 months later by a 4-mm overlapping biopsy. The collateral sprouting distance was measured. RESULTS: The mean rate of regenerative sprouting was highest for healthy control subjects (0.17 +/- 0.073 fibers/mm/day), followed by HIV+ subjects without neuropathy (0.13 +/- 0.06) and then HIV+ subjects with neuropathy (0.097 +/- 0.07) (p = 0.002). Regenerative sprouting was significantly associated with the baseline intraepidermal nerve fiber density (p < 0.001) but not with HIV viral load, HIV duration, CD4 cell count, or ApoE epsilon4 status. HIV+ individuals were also found to have a reduced collateral sprouting rate compared to controls (5.31 mum +/- 0.73 vs 9.78 mum +/- 1.5/day, p = 0.03). CONCLUSIONS: Abnormalities in both regenerative and collateral sprouting are present in people infected with HIV, and are detectable in subjects with and without evidence of peripheral nerve dysfunction. Our results indicate that abnormalities in nerve regeneration occur early in HIV infection and provide a rationale to include neuropathy-free HIV subjects in regenerative peripheral nerve trials.

Pressure-induced formation of diblock copolymer "micelles" in supercritical fluids. A combined study by small angle scattering experiments and mean-field theory. I. The critical micellization density concept.

We developed a simple mean-field theory to describe polymer and AB diblock copolymer phase separation in supercritical (SC) fluids. The highly compressible SC fluid has been described by using a phenomenological hole theory, properly extended to consider the solvent/polymer/vacancy pseudoternary mixture. The model has been applied to describe the phase behavior of AB-diblock copolymers under the assumption of a strong solvent selectivity for just one copolymer chain. In our model the solvent selectivity is a strong function of the external pressure because in compressible fluids vacancies reduce the number of favorable solvent-polymer contacts. The combined effect of the pressure on the average solvent quality and selectivity for a single polymer chain makes the phase behavior of a diblock copolymer in SC fluids quite complex. Small angle neutron and x-ray scattering (SANS and SAXS) measurements have been performed on SC-CO2 solutions of different AB-diblock copolymers containing a perfluorinated chain. The data obtained over a wide range of pressure and temperature confirm our theoretical predictions.

Pressure-induced formation of diblock copolymer "micelles" in supercritical fluids. A combined study by small angle scattering experiments and mean-field theory. II. Kinetics of the unimer-aggregate transition.

We developed a simple time-dependent mean-field theory to describe the phase separation kinetics of either homopolymers or AB-diblock copolymers in supercritical (SC) fluids. The model, previously used to describe the phase behavior of AB-block copolymers under the assumption of strong solvent selectivity for just one copolymer chain, has been extended to study the kinetics of the phase separation process. Time resolved small angle x-ray scattering (TR-SAXS) measurements have been performed on different AB-diblock copolymers containing a perfluorinated chain and dissolved in SC-CO2. The data obtained over a wide range of pressure and temperature confirm our theoretical predictions. Particularly interesting is the presence of two relaxation frequencies for the homogeneous solution --> spherical aggregate transition, where the two relaxation processes depend on the depth of the pressure jump and on temperature. The whole phenomenon could be explained as an initial SC solvent/polymer phase separation followed by a slow reorientation process to form spherical aggregates driven by the copolymer solvophilic moiety.

SAXS investigation on aggregation phenomena in supercritical CO2.

Synchrotron Small-Angle X-Ray scattering (SAXS) measurements on aggregate formation of a Polyvinyl acetate- b-Perfluoro octyl acrylate (PVAc- b-PFOA) block copolymer in supercritical CO(2) are here reported. Experiments were carried out for a series of different thermodynamic conditions, changing the solvent density by profiling both the pressure at constant temperature and the temperature at constant pressure. This block copolymer and in general fluorocarbon-hydrocarbon di-blocks form aggregates depending on the value of CO(2) density. A sharp transition between monomers dissolved as random coils and micelles characterized by a solvophilic shell and a solvophobic core occurs when the CO(2) density reaches a critical value. Results of critical micellization density (CMD) derived from pressure and temperature ramps experiment along with the comparison with previous SANS results are here reported to give additional experimental support to the solvent density-driven aggregation process.

Mass spectrometry and combinatorial chemistry: a short outline.

The rapid evolution of combinatorial chemistry in recent years has led to a dramatic improvement in synthetic capabilities. The goal is to accelerate the discovery of molecules showing affinity against a target, such as an enzyme or a receptor, through the simultaneous synthesis of a great number of structurally diverse compounds. This is done by generating combinatorial libraries containing as many as hundreds or thousands of compounds. The need to test all these compounds led to the development of high-throughput screening (HTS) techniques, and also high-throughput analytical techniques capable of assessing the occurrence, structure and purity of the products. In order to be applied effectively to the characterization of combinatorial libraries, an analytical technique must be adequately sensitive (to analyse samples which are typically produced in nanomole amounts or less), fast, affordable and easy to automate (to minimize analysis time and operator intervention). Although no method alone can meet all the analytical challenges underlying this task, the recent progress in mass spectrometric (MS) instrumentation renders this technique an essential tool for scientists working in this area. We describe here relevant aspects of the use of MS in combinatorial technologies, such as current methods of characterization, purification and screening of libraries. Some examples from our laboratory deal with the analysis of pooled oligomeric libraries containing n x 324(n = 1, 2) compounds, using both on-line high-performance liquid chromatography/MS with an ion trap mass spectrometer, and direct infusion into a triple quadrupole instrument. In the first approach, MS and product ion MS/MS with automatic selection of the precursor were performed in one run, allowing library confirmation and structural elucidation of unexpected by-products. The second approach used MS scans to characterize the entire library and also precursor ion and neutral loss scans to detect selectively components with given structural characteristics.

Structure of diblock copolymers in supercritical carbon dioxide and critical micellization pressure

This paper reports a small angle neutron scattering investigation of micelle formation by fluorocarbon-hydrocarbon block copolymers in supercritical CO2(sc-CO2) at 65 degrees C. A sharp unimer-micelle transition is obtained due to the tuning of the solvating ability of sc-CO2 by profiling pressure, so that the block copolymer, in a semidilute solution, finds sc-CO2 a good solvent at high pressure and a poor solvent at low pressure. At high pressure the copolymer is in a monomeric state with a random coil structure. However, on lowering the pressure, aggregates are formed with a structure similar to aqueous micelles with the hydrocarbon segments forming the core and the fluorocarbon segments forming the corona of the micelle. This unimer-aggregate transition is driven by the gradual elimination of CO2 molecules solvating the hydrocarbon segments of the polymer. Comparison of these results with related data on the same polymer at different temperatures indicates that the transition is critically related to the density of the solvent. This suggests the definition of a critical micellization density, to our knowledge a new concept in colloid chemistry.

Critical micellization density: A small-angle-scattering structural study of the monomer-aggregate transition of block copolymers in supercritical CO2

In this paper we report a small-angle neutron-scattering investigation of micelle formation by the fluorocarbon-hydrocarbon block copolymer, polyvinyl acetate-b-poly (1,1,2, 2-tetrahydroperfluoro-octyl acrylate) in supercritical CO2 (scCO(2)) at 313 K. At high pressure the copolymer is in a monomeric state with a random coil structure, while at low pressure the polymer forms spherical aggregates stable in a wide range of thermodynamic conditions. By profiling pressure, a sharp monomer-micelle transition is obtained due to the tuning of the solvating ability of scCO(2). We confirm the previous finding that this aggregate-monomer transition is driven by the gradual penetration of CO2 molecules toward the core of the aggregate and is critically related to the density of the solvent, thus giving additional support to the concept of a critical micellization density reported earlier on a similar polymer.

Liquid chromatography tandem mass spectrometric quantitation of sulfamethazine and its metabolites: direct analysis of swine urine by triple quadrupole and by ion trap mass spectrometry.

This work describes a new method for the quantitation of trace amounts of sulfamethazine (SMZ) and its main metabolite, N4-acetylsulfamethazine (Ac-SMZ), in swine urine, using high-performance liquid chromatography (HPLC) tandem mass spectrometric analysis of crude urine after addition of internal standard and simple dilution with water. The aim was to determine whether residues of this sulfamidic drug, normally administered to swine in order to prevent infectious diseases, were present in urine at levels lower than those permitted by regulatory authorities before human consumption (EU Project SMT, contract number CT 96-2092). A 10 microL volume of diluted urine was injected into a very short, narrow-bore chromatographic column (Zorbax SB-C18 2.1 i. d. x30 mm length, 3.5 microm pore size). Elution of the analytes of interest was achieved in less than seven minutes using a rapid gradient (from 20 to 80% methanol in 3 minutes). Either a PE Sciex API 365 triple quadrupole (QqQ), operated in the selected reaction monitoring (SRM) mode, or a Finnigan LCQ ion trap (IT) mass spectrometer, operated in narrow-range product ion scan, was used as the final detector. Electrospray (ESI) was used as the ionization technique. A comparison of the two tandem mass spectrometers was performed by analyzing the same set of test samples, at three concentration levels, on three different days. Linearity of responses of the calibration standards, intra- and inter-assay precision of the samples, specificity and limits of detection were evaluated for both systems. Both the QqQ and the IT instrument was suitable for rapid, sensitive and specific determination of the analytes, although the overall performance of the QqQ was slightly superior in terms of linearity, precision and sensitivity.

A galpha(s) carboxyl-terminal peptide prevents G(s) activation by the A(2A) adenosine receptor.

The molecular mechanisms of interaction between G(s) and the A(2A) adenosine receptor were investigated using synthetic peptides corresponding to various segments of the Galpha(s) carboxyl terminus. Synthetic peptides were tested for their ability to modulate binding of a selective radiolabeled agonist, [(3)H]2-[4-(2-carboxyethyl)phenylethylamino]-5'-N-ethylcarboxam idoade nosine ([(3)H]CGS21680), to A(2A) adenosine receptors in rat striatal membranes. The Galpha(s) peptides stimulated specific binding both in the presence and absence of 100 microM guanosine-5'-O-(3-thiotriphosphate) (GTPgammaS). Three peptides, Galpha(s)(378-394)C(379)A, Galpha(s)(376-394)C(379)A, and Galpha(s)(374-394)C(379)A, were the most effective. In the presence of GTPgammaS, peptide Galpha(s)(374-394)C(379)A increased specific binding in a dose-dependent fashion. However, the peptide did not stabilize the high-affinity state of the A(2A) adenosine receptor for [(3)H]CGS21680. Binding assays with a radiolabeled selective antagonist, [(3)H]5-amino-7-(2-phenylethyl)-2-(2-furyl)pyrazolo[4, 3-e]-1,2,4-triazolo[1,5-c]pyrimidine ([(3)H]SCH58261), showed that the addition of the Galpha(s) peptide modified the slope of the 5'-N-ethylcarboxamidoadenosine (NECA) competition curve, suggesting modulation of receptor affinity states. In the presence of GTPgammaS, the displacement curve was right-shifted, whereas the addition of Galpha(s)(374-394)C(379)A caused a partial left-shift. Both curves were fitted by one-site models. This same Galpha(s) peptide was also able to disrupt G(s)-coupled signal transduction as indicated by inhibition of the A(2A) receptor-stimulated adenylyl cyclase activity without affecting either basal or forskolin-stimulated enzymatic activity in the same membrane preparations. Shorter peptides from Galpha(s) and Galpha(i1/2) carboxyl termini were not effective. NMR spectroscopy showed the strong propensity of peptide Galpha(s)(374-394)C(379)A to assume a compact carboxyl-terminal alpha-helical conformation in solution. Overall, our results point out the conformation requirement of Galpha(s) carboxyl-terminal peptides to modulate agonist binding to rat A(2A) adenosine receptors and disrupt signal transduction.

Solid-phase synthesis of MEN 11270, a new cyclic peptide kinin B2 receptor antagonist.

An efficient synthesis of the cyclic decapeptide MEN 11270 [H-DArg1-Arg2 Pro3-Hyp4-Gly5-Thi6-Dab7-DTic8-Oic9-Arg10 c(7gamma - 10alpha)] was developed. Two three-dimensional orthogonal strategies were applied and compared: Fmoc/Tos/Boc (procedure A) and Fmoc/Pmc/Dde (procedure B). Both resulted in a 23-step strategy comprising the stepwise solid-phase chain assembly of the linear protected peptide, partial deprotection, solution-phase cyclization and final full deprotection. The stepwise assembly of the linear peptide was optimized by double coupling and acylation time prolongation for critical residues (Tic, Dab, Thi, Pro). O-(7-azabenzotriazol-1-yl)-N,N,N',N' tetramethyluronium (HATU) was preferred as coupling reagent for Dab. In the cyclization step, the partial racemization of Arg10 (31% using 1-ethyl-3-(3'-dimethyl-aminopropyl) carbodiimide/1-hydroxybenzotriazole (EDC/HOBt) as activation system) was reduced to 3% with HATU. The final deprotection was performed in the presence of dimethylsulfide (procedure A) and thiocresol (procedure B) as scavengers, to avoid the sulfation of Hyp side chain. The final compound and the main by-products were characterized by mass spectroscopy (MS), nuclear magnetic resonance (NMR) and racemization test. Procedure B produced operationally simpler and more efficient results than A (28% overall yield versus 4%).

Synthesis and biological activity of the penem antibiotic MEN 10700 and its orally absorbed ester MEN 11505.

The synthesis and biological properties of the new penem antibiotic MEN 10700 (6) and of its selected oral prodrug MEN 11505 (8f) are described. MEN 10700 showed a broad spectrum of activity, with high potency both on Gram-positive and Gram-negative strains. It also exhibited good antibacterial activity toward anaerobes and on strains selected for their resistance to other antibacterial agents (cefotaxime- or ceftazidime-resistant Gram-negative strains, ciprofloxacin-resistant E. coli, extended spectrum beta-lactamase producing and cephalosporinase inducible enterobacteria). MEN 10700 showed a very high stability to enzymatic degradation by renal dehydropeptidase DHP-I. After oral administration in rats of the pivaloyloxymethyl ester prodrug MEN 11505, the relative bioavailability of MEN 10700 was calculated as F=43%.

A new class of pseudopeptide antagonists of the kinin B1 receptor containing alkyl spacers.

Four previously reported kinin receptor peptide antagonists, including the B1 receptor-selective peptides desArg10-HOE 140 (H-D-Arg-Arg-Pro-Hyp-Gly-Thi-Ser-D-Tic-Oic-OH) and B-9858 (H-Lys-Lys-Arg-Pro-Hyp-Gly-Igl-Ser-D-Igl-Oic-OH), have been modified by replacement of the central tetrapeptide Pro-Hyp-Gly-Xaa with linear alkyl spacers of variable length. The analogue of desArg10-HOE 140 containing the 11-aminoundecanoic acid as spacer, MEN 11575 [H-D-Arg-Arg-NH-(CH2)10-CO-Ser-D-Tic-Oic-OH], was found to be slightly more potent than the unmodified peptide (pA2 = 7.1) as a kinin B1 receptor antagonist in the rat ileum longitudinal smooth muscle assay. Moreover, MEN 11575 is devoid of residual agonist activity at the kinin B1 receptor (rat ileum) and antagonist activity at the kinin B2 receptor (guinea pig ileum longitudinal smooth muscle). Both these activities are displayed by the parent peptide desArg10-HOE 140. Therefore, despite its greatly simplified chemical structure, MEN 11575 shows an improved pharmacological profile in terms of both potency and selectivity, and it represents a good template for the development of new peptidomimetic kinin B1 receptor antagonists. We also report an attempt to investigate the conformational role of the flexible, linear spacer of MEN 11575 and to design more constrained analogues, possibly locked in the bioactive conformation, using semirigid spacers based on Calpha-tetrasubstituted alpha-amino acids of the family of 1-aminocycloalkane-1-carboxylic acids (Acnc).

Non-covalent complexes between DNA-binding drugs and double-stranded deoxyoligonucleotides: a study by ionspray mass spectrometry.

The non-covalent complexes between some DNA-binding drugs and duplex oligodeoxynucleotides were studied by ionspray mass spectrometry, with the aim of evaluating the suitability of this technique to screen rapidly a series of drugs exerting their activity through non-covalent binding to specific base sequences of DNA. Two classes of drugs were considered, distamycins (which show affinity for the minor groove of DNA) and anthracyclines (which interact through intercalation between bases). For the former, d(CGCGAATTCGCG)2 was chosen as the model oligodeoxynucleotide. Following optimization of sample preparation and instrumental conditions, the complexes of different distamycins were observed; depending on the ligand considered, 1:1 or 2:1 complexes were formed preferentially. A semi-quantitative evaluation of the relative affinities was made by measuring the ratio of the complexes signals to those of the duplex, and also by competitive binding with equimolar amounts of distamycin. For anthracyclines, the daunorubicin-d(CGATCG)2 complex was chosen as the model for a preliminary mass spectrometric study; however, the signals of the duplex and the complex were very low compared with the monomer signal. Since the complex was known to be stable in solution, this was ascribed to gas-phase instability, probably caused by electrostatic repulsion between negatively charged phosphate groups.