Carbon nanotube/polysulfone soft composites: preparation, characterization and application for electrochemical sensing of biomarkers.

Multi-walled carbon nanotube/polysulfone soft composites (MWCNT/PSf) prepared via phase inversion are a novel platform for electrochemical and electroanalytical purposes with practical applications in the design of screen-printed electrodes for electrochemical sensing. We present here a thorough characterization of the morphological, physical, chemical and electrochemical properties of this material. These composites constitute a robust mesoporous network with high specific surface area, which is beneficial for trapping bioanalytes and increasing the electrochemical sensitivity. We highlight the advantages of these soft composites by comparing them with analogous graphite composites.

Electrocatalyzed O2 response of myoglobin immobilized on multi-walled carbon nanotube forest electrodes.

Direct electrochemistry and activity of myoglobin (Mb) immobilized on carbon nanotube (CNT) forest electrodes were investigated by probing mainly its electrocatalytical response towards oxygen. The protein was anchored on the CNT electrodes through carbodiimide coupling, which was shown to provide long term stability. The electrochemical response was monitored as a function of oxygen concentration and pH. Conformational changes together with the consequent loss of oxygen affinity were recorded at low pH, which delimits the operative range of the Mb/CNT electrodes for sensing purposes. In general, it can be concluded that CNT forests constitute suitable platforms for Mb attachment without compromising the protein bioactivity and by keeping at the same time the direct electron exchange with the heme core. All these characteristics confer to the protein modified CNT system promising properties for the implementation of (bio)sensor devices with impact in the clinical and environmental field.

Enhancing the electrochemical response of myoglobin with carbon nanotube electrodes.

In this paper, the electrochemical behavior of different myoglobin-modified carbon electrodes is evaluated. In particular, the performance of voltammetric biosensors made of forest-like carbon nanotubes, carbon nanotube composites and graphite composites is compared by monitoring mainly the electrocatalytic reduction of H(2)O(2) by myoglobin and their corresponding electroanalytical characteristics. Graphite composites showed the worst electroanalytical performance, exhibiting a small linear range, a limit of detection (LOD) of 9 x 10(-5) M and low sensitivity. However, it was found that the electrochemical response was enhanced with the use of carbon nanotube-based electrodes with LOD up to 5 x 10(-8) M, higher sensitivities and wider linear range response. On the one hand, in the case of the CNT epoxy composite, the improvement in the response can be mainly attributed to its more porous surface which allows the immobilization of higher amounts of the electroactive protein. On the other hand, in the case of the forest-like CNT electrodes, the enhancement is due to an increase in the electron transfer kinetics. These findings encourage the use of myoglobin-modified carbon nanotube electrodes as potential (bio)sensors of H(2)O(2) or O(2) in biology, microbiology and environmental fields.

Residual aqueous ozone determination by gas diffusion reverse flow injection analysis.

A novel system based on reverse flow injection analysis with a gaseous diffusion step (GD-r-FIA) has been developed for the analysis of ozone. It includes an automatic microburet injection system. The ozone diffuses through a microporous membrane of polyvinylidene difluoride (PVDF) from the donor stream to the acceptor stream containing nitrite ions. The nitrite concentration in the acceptor solution decreases due to the ozone reduction reaction. In this way, a simple indirect measurement of the ozone concentration can be performed using the Griess-Ilosvay reaction for the nitrite ion. This correlates with the decrease in absorbance of the azoic dye formed with the ozone concentration in the donor stream. The system has been optimised by investigating the effect of the nitrite concentration in the acceptor stream on the diffusion flow. The optimum nitrite concentration was set at 0.250 ppm with a flow rate of 1.5 ml/min. The efficiency of the ozone diffusion through the membrane was only 4.4%. This affects the average sensitivity, which is low (0.0092+/-0.0012 AU/ppm), although the detection limit is similar to that obtained with other reported methods (0.03 ppm). The main advantage of the system reported here is that it has a linear range that is an order of magnitude broader than those observed for other GD-FIA systems. This is especially useful for continuous monitoring systems, since the residual ozone concentration is normally between 0.05 and 5.0 ppm. Additionally, using the reverse flow injection analysis (FIA) technique minimises chemical consumption and residue generation. Finally, the stability of the ozone solution and the repeatability and reproducibility of the method have been studied.

Autoadaptative sequential injection system for nitrite determination in wastewaters.

A novel autoadaptative sequential injection system for the analysis of nitrite is described. The automatic determination uses a direct spectrophotometric method, based on the Griess-Ilosvay reaction. In this method the absorbance of the purple azo dye formed is measured at 555nm. In the sequential injection operation, the sample and the reagent are aspirated and mixed by reverse flow. The sequencing and overlapping of stacked (reagent) zones as well as selection of volumes have been studied in detail. The proposed analytical system is intelligent, simple and robust, allowing for nitrite determination in a double concentration range, by a simple and automatic programmable operation change. These two ranges are 0.0-3.0 and 0.0-20.0ppm with detection limits being 0.048 and 0.4ppm, respectively. Next surroundings have been developed allowing autocalibration and independent monitoring of nitrite concentration. The experimental set-up has been evaluated applying it to real samples analysis of very diverse concentration samples coming from a WWTP. The throughput of the method was 12 samples per hour.

In vitro and in vivo efficacies of the new triazole albaconazole against Cryptococcus neoformans.

The activity of albaconazole (UR-9825; J. Uriach & Cía. S.A., Barcelona, Spain) was compared to that of fluconazole against 12 isolates of Cryptococcus neoformans in vitro and against 1 isolate in vivo in a rabbit model of cryptococcal meningitis. Albaconazole was 100-fold more potent in vitro than fluconazole on a per-weight basis and was fungicidal at potentially relevant concentrations for two isolates. MICs ranged from

Issues in system boundary definition for substance flow analysis: the case of nitrogen cycle management in Catalonia.

The great complexity of the nitrogen cycle, including anthropogenic contributions, makes it necessary to carry out local studies, which allow us to identify the specific cause-effect links in a particular society. Models of local societies that are based on methods such as Substance Flow Analysis (SFA), which study and characterise the performance of metabolic exchanges between human society and the environment, are a useful tools for directing local policy towards sustainable management of the nitrogen cycle. In this paper, the selection of geographical boundaries for SFA application is discussed. Data availability and accuracy, and the possibility of linking the results with instructions for decision making, are critical aspects for proper scale selection. The experience obtained in the construction of the model for Catalonia is used to draw attention to the difficulties found in regional studies.

In vitro antiproliferative effects and mechanism of action of the new triazole derivative UR-9825 against the protozoan parasite Trypanosoma (Schizotrypanum) cruzi.

We describe the in vitro antiproliferative effects of the new triazole derivative UR-9825 against the protozoan parasite Trypanosoma (Schizotrypanum) cruzi, the causative agent of Chagas' disease in Latin America. The compound was found to be extremely active against the cultured (epimastigote) form of the parasite, equivalent to that present in the reduviid vector, with a MIC of 30 nM, a concentration 33-fold lower than that required with the reference compound ketoconazole. At that MIC, growth arrest coincided with depletion of the parasite's 4,14-desmethyl endogenous sterols (ergosterol, 24-ethylcholesta-5,7,22-trien-3b-ol, and precursors) and their replacement by methylated sterols (lanosterol, 24-methylenedihydrolanosterol, and obtusifoliol), as revealed by high-resolution gas chromatography coupled with mass spectrometry. This indicated that the primary mechanism of action of UR-9825 was inhibition of the parasite's sterol C14alpha demethylase, as seen with other azole derivatives. The phospholipid composition of growth-arrested epimastigotes was also altered, when compared to controls, with a significant increase in the content of phosphatidylethanolamine and phosphatidylserine and a concomitant reduction of the content of phosphatidylcholine. The clinically relevant intracellular amastigote form, grown in cultured Vero cells at 37 degrees C, was even more sensitive to UR-9825, with a MIC of 10 nM, comparable to that for ketoconazole. The results showed that UR-9825 is among the most potent azole derivatives tested against this parasite and support in vivo studies with this compound.

New azole antifungals. 2. Synthesis and antifungal activity of heterocyclecarboxamide derivatives of 3-amino-2-aryl-1-azolyl-2-butanol.

A series of 92 azole antifungals containing an amido alcohol unit was synthesized. The nature and substitution of the amide portion was systematically modified in search of improved antifungal activity, especially against filamentous fungi. The compounds were tested in vitro against a variety of clinically important pathogens and in vivo (po) in a murine candidosis model. Thiazole and thiophene carboxamides carrying both a substituted phenyl ring and a small alkyl group were best suited for activity against filamentous fungi. In a subset of these compounds, the amide portion was conformationally locked by means of a pyrimidone ring and it was proven that only an orthogonal orientation of the phenyl ring yields bioactive products. A tendency to display long plasma elimination half-lives was observed in both series. Two compounds, 74 and 107, representative of the open and cyclic amides, respectively, were chosen for further studies, based on their excellent activity in in vivo murine models of candidosis and aspergillosis. This work describes the SARs found within this series. The next paper displays the results obtained in a related series of compounds, the quinazolinones.

New azole antifungals. 3. Synthesis and antifungal activity of 3-substituted-4(3H)-quinazolinones.

A series of azole antifungal agents featuring a quinazolinone nucleus have been subjected to studies of structure-activity relationships. In general, these compounds displayed higher in vitro activities against filamentous fungi and shorter half-lives than the structures described in our preceding paper. The most potent products in vitro carried a halogen (or an isostere) at the 7-position of the quinazolinone ring. Using a murine model of systemic candidosis, oral activity was found to be dependent on hydrophobicity, which, in turn, modulated the compound's half-life. The 7-Cl derivative, (1R,2R)-7-chloro-3-[2-(2, 4-difluorophenyl)-2-hydroxy-1-methyl-3-(1H-1,2, 4-triazol-1-yl)propyl]quinazolin-4(3H)-one (20, UR-9825), was selected for further testing due to its high in vitro activity, low toxicity, good pharmacokinetic profile, and ease of obtention. Compound 20 is the (1R,2R) isomer of four possible stereoisomers. The other three isomers were also prepared and tested. The enantiomer (1S,2S) and the (1R,2S) epimer were inactive, whereas the (1S,2R) epimer retained some activity. In vitro 20 was superior to fluconazole, itraconazole, SCH-42427, and TAK-187 and roughly similar to voriconazole and ER-30346. In vivo, 20 was only moderately active in a mouse model of systemic candidosis when administration was limited to the first day. This was attributed to its short half-life in that species (t1/2 = 1 h po). Protection levels comparable to or higher than those of fluconazole, however, were observed in systemic candidosis models in rat and rabbit, where the half-life of the compound was found to be 6 and 9 h, respectively. Finally, 20 showed excellent protection levels in an immunocompromised rat model of disseminated aspergillosis. The compound showed low toxicity signs when administered to rats at 250 mg/kg qd or at 100 mg/kg bid during 28 days.

Photosensitive polyurethanes applied to the development of CHEMFET and ENFET devices for biomedical sensing.

Chemical microsensors based on ion-selective field effect transistor (ISFET) transducers with ion-selective and enzymatic membranes have been fabricated. In this case, photolithographically patterned membranes based on acrylated urethanes have been developed and applied onto the gate area of ISFET chips. Aliphatic urethane diacrylate has been used for K+ and NH+4 membranes, while a photocurable hydrogel formulation based on other type of acrylated urethane has been optimized for urea-FET sensors. Resulting potassium and ammonium sensors show similar performances to those found when PVC membranes are employed. An integrated packaging process for ISFET-based sensors has been developed giving the possibility of carrying out most of the encapsulation on wafer level. For this purpose, a photocurable polyurethane encapsulant formulation has been optimized to be microstructured by photolithography. Finally, a preliminary study of biocompatibility of photosensitive formulations containing urethane oligomers has been performed in order to examine future applications in biomedical and clinical analysis.

Continuous-Flow System for On-Line Water Monitoring Using Back-Side Contact ISFET-Based Sensors.

A multiparametric continuous-flow system for on-line monitoring of water based on ISFET sensors is described. The ISFETs used have silicon nitride as gate material, and the electrical contacts are placed on the back side of the chip. This is a technological improvement that allows for a more compact ISFET packaging and greatly increases the lifetime of the sensor compared with planar type ISFETs, since the electrical parts are separated from the chemical environment. A special probe has been designed in order to encapsulate and apply these ISFETs into the flow system. Further, a reference electrode based on standard Ag/AgCl technology has been constructed according to the ISFET probe design in order to integrate both sensors in the same flow-through cell. These probes can be easily replaced in the flow system and are made of cheap and easily mechanized materials. Using these flow-through sensors, a continuous-flow system for the determination of pH, NH(4)(+), Ca(2+), and NO(3)(-) in waters has been designed. The system configuration is based on a modular design (one setup for each parameter and a common sampling channel), which allows simple manipulation and maintenance as well as a good flexibility for different analytical requirements. A study of the system characteristics was performed by following the specifications for water monitoring. Under the conditions established for the flow system, a sampling rate of 20 h(-)(1) was obtained for each parameter, and long-term stabilities of at least 3 weeks of daily work for ISFET sensors and 5 months for the reference electrode have been achieved. The response performances obtained show the feasibility of the BSC ISFET probe use in continuous-flow monitoring.

Synthesis and antifungal activity of new azole derivatives containing an N-acylmorpholine ring.

A series of azole derivatives carrying an N-acylmorpholine ring are described. The compounds were chemically designed to simulate the lanosterol D ring, taking advantage of the conformational preferences of 2-alkyl-1-acylmorpholines. Three structural variables, the nature of the N-benzoyl group, the phenyl substituents, and the degree of oxidation at carbon 2 of the morpholine, were optimized for maximum activity. Only the (5R,6R) isomers showed antifungal activity. Cyclic hemiacetal (-)-39a (UR-9746) and cyclic ether (-)-41 (UR-9751) were selected for further development. In vitro, (-)-41 was clearly more active than (-)-39a and somewhat less active than the acyclic counterpart (-)-7. In vivo activity was assessed by a systemic (mouse) and a vaginal (rat) candidosis model. In the former, (-)-39a, (-)-41, and (-)-7 at 1 mg/kg given 1, 4, and 24 h postinfection displayed 90-100% protection from mortality on day 9. Compound (-)-39a was slightly more potent than (-)-41 and similar in potency to (-)-7. The three compounds were superior in potency to fluconazole and similar in potency in SCH-42427 in this test. In the vaginal model, (-)-39a and (-)-41 given daily during 3 days after infection at 0.5 mg/kg showed high levels of protection on days 10 and 15. At 0.25 mg/kg, (-)-39a was slightly more potent than SCH-42427 and (-)-7 and superior in potency to (-)-41 and fluconazole in this model. Preliminary 28-day toxicity tests at 100 mg/kg/day po in rats indicated no or very mild adverse effects for the two UR compounds.

[(3-Pyridylalkyl)piperidylidene]benzocycloheptapyridine derivatives as dual antagonists of PAF and histamine.

A series of [(3-pyridylalkyl)piperidylidene]- and (nicotinoylpiperidylidene)benzocycloheptapyridine derivatives, Ia,b, were prepared and evaluated for PAF antagonist and H1 antihistamine activity. PAF antagonist activity was investigated by the in vitro PAF-induced platelet aggregation assay (PPA) and the in vivo PAF-induced hypotension test in rats (PH) and mortality test in mice (PM). For the evaluation of H1 antihistamine activity, the in vitro histamine-induced contraction of the guinea-pig ileum assay (HC) and the in vivo histamine-induced hypotension test (HH) in normotensive rats were used. The potential antiallergic activity of the compounds was evaluated using the active anaphylactic shock test in mice. These compounds are structurally related to loratadine (1) and were generated by replacement of the ethoxycarbonyl group of 1 with substituted 3-pyridylmethyl and nicotinoyl moieties. Both anti-PAF and H1 antihistamine activities have shown a high dependence on the exact nature and position of the substituent in the pyridine ring. Optimum structure 19 (UR-12592) incorporating a (5-methyl-3-pyridyl)methyl radical displayed an unique dual activity inhibiting both PAF-induced effects (PPA, IC50 = 3.7 microM; PH, ID50 = 0.44 mg/kg iv; PM, ID50 = 1.9 mg/kg po) and histamine-induced effects (HC, IC50 = 3.9 nM; HH, ID50 = 1.4 mg/kg iv). Furthermore, 19 was highly active in the passive cutaneous anaphylactic shock in rats (ID50 = 1.2 mg/kg po) and strongly protected mice and rats from mortality induced by endotoxin (ID50 = 1.2 and 0.5 mg/kg iv, respectively). Compound 19 showed itself to be devoid of CNS depressant effects, neither modifying spontaneous motor activity nor prolonging barbiturate-sleeping time in mice at a dose of 100 mg/kg po, and is now under development.

Synthesis and structure-activity relationships of 1-acyl-4-((2-methyl-3-pyridyl)cyanomethyl)piperazines as PAF antagonists.

A second generation of (cyanomethyl)piperazines, 1-acyl-4-((2-methyl-3-pyridyl)cyanomethyl)-piperazines, with increased oral activity was prepared and evaluated in vitro in a PAF-induced platelet aggregation assay (PAG) and in vivo in a PAF-induced hypotension test in normotensive rats (HYP). Oral activity was ascertained through a PAF-induced mortality test in mice (MOR). Attachment of a methyl group at position 2 of our earlier pyridine derivatives resulted in an improvement of 1 order of magnitude or greater in the ID50 of the oral test. Three different types of acyl substituents of similar potency emerge from this work: N-(diphenylmethylamino)acetyl, 3-substituted 3-hydroxy-3-phenylpropionyl, and N-substituted 3-amino-3-phenylpropionyl groups. The most interesting compounds, 26 (UR-12460, PAG IC50 = 0.040 microM, HYP, ID50 = 0.021 mg/kg i.v., MOR, ID50 = 0.30 mg/kg po) and 58 (UR-12519, PAG IC50 = 0.041 microM, HYP, ID50 = 0.015 mg/kg i.v., MOR, ID50 = 0.044 mg/kg po), compare favorably with WEB-2086. Compounds 26 and 58 were also tested in active anaphylactic shock (AAS) and endotoxin-induced mortality (EIM) tests. On the basis of these data, compounds 26 and 58 have been selected for further pharmacological development.

Comparison of the simplex and Powell methods with a weighted response function for the optimization of FIA systems.

An experimental comparison between the relative performances of the Powell and simplex methods for the optimization of a Flow Injection Analysis (FIA) system for the determination of nitrite in water is reported. An evaluation of the advantages of using a weighted linear combination of two variables (related to sensitivity and sample throughput rate) as the response function in the guidance of the optimization procedures towards different practical requirements is also included. Both methods proved to be effective for the optimization, none having shown definite advantages over the other. The use of a weighted response function in these optimization methods proved to be useful for assessing the versatility of FIA systems.

2,2-Dialkylnaphthalen-1-ones as new potassium channel activators.

A new series of 2,2-dialkylnaphthalen-1-one potassium channel activators has been prepared, and their in vitro relaxant activities in isolated rat portal vein and guinea pig tracheal spirals as well as their oral antihypertensive effect in spontaneously hypertensive rats have been evaluated. The group of 1,2-dihydro-4-(1,2-dihydro-2-oxo-1-pyridyl)-2,2-dimethylnaphthalen -1- ones with an electron-withdrawing substituent at the 6-position contain the most active compounds and 1,2-dihydro-4-(1,2-dihydro-2-oxo-1-pyridyl)-2,2-dimethyl-1-oxonaphtha lene-6- carbonitrile, 17f (UR-8225), has been selected for further pharmacological development.

(Pyridylcyanomethyl)piperazines as orally active PAF antagonists.

A series of (pyridylcyanomethyl)piperazines was prepared and evaluated for PAF-antagonist activity. Compounds were tested in vitro in a PAF-induced platelet aggregation assay and in vivo in a PAF-induced hypotension test in normotensive rats. Oral activity was ascertained through a PAF-induced mortality test in mice. The main structure-activity trends of the series were established. Activity was mainly found in four skeletons: 1-acyl-4-(3-pyridylcyanomethyl)piperazine, 1-acyl-4-(4-pyridylcyanomethyl)piperazine, 1-acyl-4-(3-pyridylcyanomethyl)piperidine, and 1-acyl-4-cyano-4-(3-pyridylamino)piperidine. The acyl substituents, diphenylacetyl and 3,3-diphenylpropionyl, provided the most active compounds, and the introduction of an amine or hydroxy group in the 3,3-diphenylpropionyl substituent led to further improvement in oral activity. As a result, three of the most active compounds (100, 114, and 115) have been selected for further pharmacological development.

4-substituted 2-alkoxytetrahydrofurans as potent and long-lasting PAF antagonists.

A series of 4-substituted 2-alkoxytetrahydrofuran derivatives featuring an acetal group were prepared and evaluated for PAF antagonist activity in the PAF-induced in vitro platelet-aggregation and in vivo hypotension tests. Compound 2-[[N-acetyl-N-[[[2-(octadecyloxy)tetrahydrofuran-4- yl]methoxy]carbonyl]amino]methyl]-1-ethylpyridinium chloride (4e, UR-11353) was selected for further development on the basis of its high activity and long-lasting action. The compound maintained a significant activity even 24 h after administration of a single dose of 1 mg/kg iv in the PAF-induced mortality test in mice and 10 h after administration of the same dose in the PAF-induced hypotension test in rats. Comparison with previously reported carba analogues suggests that the presence of the acetal group is the structural characteristic that confers its long-lasting activity.