Simultaneous determination of rosiglitazone and its metabolites in rat liver microsomal fraction using hollow-fiber liquid-phase microextraction for sample preparation

A three-phase hollow-fiber liquid-phase microextraction method for the analysis of rosiglitazone and its metabolites N-desmethyl rosiglitazone and p-hydroxy rosiglitazone in microsomal preparations is described for the first time. The drug and metabolites HPLC determination was carried out using an X-Terra RP-18 column, at 22 degrees C. The mobile phase was composed of water, acetonitrile and acetic acid (85:15:0.5, v/v/v) and the detection was performed at 245 nm. The hollow-fiber liquid-phase microextraction procedure was optimized using multifactorial experiments and the following optimal condition was established: sample agitation at 1750 rpm, extraction for 30 min, hydrochloric acid 0.01 mol/L as acceptor phase, 1-octanol as organic phase, and donor phase pH adjustment to 8.0. The recovery rates, obtained by using 1 mL of microsomal preparation, were 47-70%. The method presented LOQs of 50 ng/mL and it was linear over the concentration range of 50-6000 ng/mL, with correlation coefficients (r) higher than 0.9960, for all analytes. The validated method was employed to study the in vitro biotransformation of rosiglitazone using rat liver microsomal fraction.

Chiral HPLC analysis of venlafaxine metabolites in rat liver microsomal preparations after LPME extraction and application to an in vitro biotransformation study

A three-phase LPME (liquid-phase microextraction) method for the enantioselective analysis of venlafaxine (VF) metabolites (O-desmethylvenlafaxine (ODV) and N-desmethylvenlafaxine (NDV) in microsomal preparations is described for the first time. The assay involves the chiral HPLC separation of drug and metabolites using a Chiralpak AD column under normal-phase mode of elution and detection at 230 nm. The LPME procedure was optimized using multifactorial experiments and the following optimal condition was established: sample agitation at 1,750 rpm, 20 min of extraction, acetic acid 0.1 mol/L as acceptor phase, 1-octanol as organic phase and donor phase pH adjustment to 10.0. Under these conditions, the mean recoveries were 41% and 42% for (-)-(R)-ODV and (+)-(S)-ODV, respectively, and 47% and 48% for (-)-( R)-NDV and (+)-( S)-NDV, respectively. The method presented quantification limits of 200 ng/mL and it was linear over the concentration range of 200-5,000 ng/mL for all analytes. The validated method was employed to study the in vitro biotransformation of VF using rat liver microsomal fraction. The results demonstrated the enantioselective biotransformation of VF.

Anti-inflammatory and antinociceptive effects of Baccharis dracunculifolia DC (Asteraceae) in different experimental models

Aim of the study: The aerial parts of Baccharis dracunculifolia D.C., popularly known as ""alecrim do campo"" are used in folk medicine as anti-inflammatory. The aim of the present study was to evaluate the anti-inflammatory and antinociceptive activities of the crude hydroalcoholic extract obtained from leaves of Baccharis dracunculifolia (BdE), which have not been reported. Matetials and methods: BdE was analyzed by HPLC and in vivo evaluated (doses ranging from 50 to 400 mg/kg, p.o.) by using the acetic acid-induced abdominal constrictions, paw oedema induced by carrageenan or histamine, overt nociception models using capsaicin, glutamate or phorbol myristate acetate (PMA), formalin-induced nociception and mechanical hypernociception induced by carrageenan or complete Freund adjuvant (CFA). As positive controls it was used paracetamol in both acetic acid and formalin tests; dipyrone in capsaicin, glutamate and PMA-induced nociception; indomethacin in CFA and carrageenan-induced hypernociception models. In addition, the in vitro effects of BdE on COX-2 activity and on the activation of NF-kappa B were also evaluated. Results: BdE (50-400 mg/kg, p.o.) significantly diminished the abdominal constrictions induced by acetic acid, glutamate and CFA. Furthermore, BdE also inhibited the nociceptive responses in both phases of formalin-induced nociception. BdE, administered orally, also produced a long-lasting anti-hypernociceptive effect in the acute model of inflammatory pain induced by carrageenan. It was also observed the inhibition of COX-2 activity by BdE. Conclusion: In summary, the data reported in this work confirmed the traditional anti-inflammatory indications of Baccharis dracunculifolia leaves and provided biological evidences that Baccharis dracunculifolia, like Brazilian green propolis, possess antinociceptive and anti-inflammatory activities. (C) 2009 Elsevier Ireland Ltd. All rights reserved.

Engineered ecosystem for sustainable on-site wastewater treatment

A Pilot-Scale Engineered Ecosystem (PSEE) operated for over two years in sub-tropical conditions, produced an effluent with COD (median 38 mg/L) and TSS (median 3 mg/L) levels comparable to that required by the AS/NZS 1547:2000 Onsite Domestic Wastewater Management standard. Only partial nitrification was achieved as dissimilatory nitrate reduction to ammonia occurred; however the level of NH4-N was reduced by 75% and total inorganic nitrogen by 53%. Phosphorus was not removed by the system due to the lack of regular sludge removal. Mass balances around the system showed that bacteria removed 36% of the influent nitrogen and 76% of the influent COD. Algae and plants were shown to remove 5% of the influent nitrogen, and 6% of the influent phosphorus. Challenges in developing a sustainable on-site wastewater treatment system were largely met by minimising chemical, energy and labour inputs, eliminating the need for frequent sludge handling, and creating an effluent quality suitable for re-use in non-potable applications. However, the sludge removal from the system needs to be adequately managed to avoid excessive accumulation as this can cause a range of negative impacts.

Vitamin D receptor expression in the embryonic rat brain

We are interested in determining whether low maternal vitamin D-3 affects brain development in utero. Whilst the vitamin D receptor (VDR) has been identified in embryonic rat brains, the timing and magnitude of its expression across the brain remains unclear. In this study we have quantitated VDR expression during development as well correlated the timing of its appearance with two vital developmental events, apoptosis and mitosis. Brains from embryonic rats (embryonic days 15-23) were examined. We show that the well-described increase in apoptotic cells and decrease in mitotic cells during development correlates with the appearance of the VDR in brain tissue. Given that vitamin D-3 regulates mitosis and apoptosis in non-neuronal tissue we speculate that the timing of VDR expression in embryonic brain may directly or indirectly mediate features of neuronal apoptosis and mitosis.

Activity of platinum-tin catalysts prepared by the Pechini-Adams method for the electrooxidation of ethanol

Pt-Sn electrocatalysts of different compositions were prepared and dispersed on carbon Vulcan XC-72 using the Pechini-Adams method. The catalysts were characterized by energy dispersive X-ray analysis and X-ray diffraction. The electrochemical properties of these electrode materials were also examined by cyclic voltammetry and chronoamperometric experiments in acid medium. The results showed that the presence of Sn greatly enhances the activity of Pt towards the electrooxidation of ethanol. Moreover, it contributes to reduce the amount of noble metal in the anode of direct alcohol fuel cells, which remains one of the challenges to make the technology of direct alcohol fuel cells possible. Electrolysis of ethanol solutions at 0.55 V vs. RHE allowed to determine by liquid chromatography acetaldehyde and acetic acid as the main reaction products. CO(2) was also analyzed after trapping it in a NaOH solution indicating that the cleavage of the C-C bond in the ethanol molecule did occur during the adsorption process. In situ IR reflectance spectroscopy helped to investigate in more details the reaction mechanism through the identification of the reaction products as well as the presence of some intermediate adsorbed species, such as linearly bonded carbon monoxide. (C) 2009 Elsevier B.V. All rights reserved.

Primidone oxidation catalyzed by metalloporphyrins and Jacobsen catalyst

Primidone (PRM) oxidation by various oxidants such as iodosylbenzene (PhIO), tert-butyl hydroperoxide 70wt.% (t-BOOH), 3-chloroperoxybenzoic acid (m-CPBA) and hydrogen peroxide 30wt.%, mediated by either a salen complex or metalloporphyrins, was investigated. The catalytic systems led to phenylethyl-malondiamide (PEMA) and phenobarbital (FEND), the same metabolites obtained in vivo with P450 enzymes, although three other products were also detected. Product formation was highly dependent on the oxidant, co-catalyst (imidazole), pH and dioxygen. These biomimetic chemical models have potential application in the synthesis of drug metabolites. which should provide samples for pharmacological tests. They can also be employed in studies that pursue the elucidation of in vivo drug metabolism. (C) 2008 Elsevier B.V. All rights reserved.

Effect of Ni on Pt/C and PtSn/C prepared by the Pechini method

Different compositions of Pt, PtNi, PtSn, and PtSnNi electrocatalysts supported on carbon Vulcan XC-72 were prepared through thermal decomposition of polymeric precursors. The nanoparticles were characterized by morphological and structural analyses (XRD, TEM, and EDX). XRD results revealed a face-centered cubic structure for platinum, and there was evidence that Ni and Sn atoms are incorporated into the Pt structure. The electrochemical investigation was carried out in slightly acidic medium (H(2)SO(4) 0.05 mol L(-1)), in the absence and in the presence of ethanol. Addition of Ni to Pt/C and PtSn/C catalysts significantly shifted the onset of ethanol and CO oxidations toward lower potentials, thus enhancing the catalytic activity, especially in the case of the ternary PtSnNi/C composition. Electrolysis of ethanol solutions at 0.4 V us. RHE allowed for determination of acetaldehyde and acetic acid as the reaction products, as detected by HPLC analysis. Due to the high concentration of ethanol employed in the electrolysis experiments (1.0 mol L(-1)), no formation of CO(2) was observed. Copyright (C) 2010, Hydrogen Energy Publications, LLC. Published by Elsevier Ltd. All rights reserved.

Tetra-crowned porphyrin as P450 biomimetic model for carbamazepine oxidation

A substituted porphyrin bearing four crown ether units, H(2)(TCP), was synthesized from the reaction between (5,10,15,20-tetra(o-aminophenyl) porphyrin) and the acyl derivative of the ether (4-carboxy-18-crown-6). The free-base porphyrin was characterized by C, N, and H elemental analysis; UV-vis and IR spectroscopies; and (1)H NMR. The corresponding ironporphyrin, Fe(TCP)Cl, was obtained via iron insertion into H(2)(TCP). Fe(TCP)Cl was employed as catalyst for carbamazepine (CBZ) oxidation by iodosylbenzene (PhIO), 3-chloroperoxybenzoic acid (m-CPBA) or sodium hypochlorite (NaOCl), in methanol or in a biphasic water/dichloroethane system. The crowned ironporphyrin proved to be a highly efficient and selective catalyst for CBZ epoxidation even in the biphasic dichloroethane /H(2)O system, with no need for an additional phase transfer agent.

Changes in purines concentration in the cerebrospinal fluid of patients experiencing pain: A case-control study

This study analyzes the relationship between extracellular purines and pain perception in humans. Cerebrospinal fluid (CSF) levels of purines and their metabolites were compared between patients displaying acute and/or chronic pain syndromes and control subjects. The CSF levels of IMP, inosine, guanosine and uric acid were significantly increased in the chronic pain group and correlated with pain severity (P<0.05). Patients displaying both chronic and acute pain presented similar changes in the CSF purines concentration (P<0.05). However, in the acute pain group, only CSF inosine and uric acid levels were significantly increased (P<0.05). These findings suggest that purines, in special inosine, guanosine and uric acid, are associated with the spinal mechanisms underlying nociception. (C) 2010 Elsevier Ireland Ltd. All rights reserved.

Markers of vascular differentiation, proliferation and tissue remodeling in juvenile nasopharyngeal angiofibromas

Juvenile nasopharingeal angiofibroma (JNA) is a histologically benign locally aggressive tumor characterized by irregular vessels embedded. in a fibrous stroma. Excessive vascularity results in bleeding complications, and the inhibition of angiogenesis is a promising strategy for managing extensive JNA tumors. To better characterize the endothelial components of JNA, we aimed to evaluate markers of vascular differentiation and proliferation, such as friend leukemia integration-1 (FLI-1) and endoglin, lymphatic markers, including podoplanin and vascular endothelial growth factor receptor 3 (VEGFR3) and its cognate ligand VEGFC, GLUT-1, a diagnostic marker that discriminates between hemangiomas and vascular malformations, and two markers of tissue remodeling, stromelysin 3 (ST3) and secreted acid protein rich in cysteine (SPARC). Antigens were assessed immunohistochemically in vessels and stromal cells of JNA archival cases (n=22). JNA endothelial cells were positive for endoglin, VEGFC and FLI-1, whereas podoplanin and VEGFR3 were negative in all cases. Both endothelial cells and fibroblasts stained for ST3 and SPARC. GLUT-1 was investigated in JNA cases, in infantile hemangiomas (n=123) and in vascular malformations (n=135) as controls. JNAs and vascular malformations were GLUT-1-negative, while hemangiomas showed positive staining. The presence of markers of endothelial differentiation and proliferation highlighted the hyper-proliferative state of JNA vessels. The absence of podoplanin and VEGFR3 underscores their blood endothelial cell characteristic. The absence of GLUT-1 discriminates JNAs from hemangiomas. ST3 and SPARC up-regulation in endothelial cells and fibroblasts may contribute to a compensatory signaling for controlling angiogenesis. Some of these markers may eventually serve as therapeutic targets. Our results may aid in the understanding of JNA pathophysiology.

Differential display and suppression subtractive hybridization analysis of the pulp of ripening banana

The fruit of banana undergoes several important physico-chemical changes during ripening. Analysis of gene expression would permit identification of important genes and regulatory elements involved in this process. Therefore, transcript profiling of preclimacteric and climacteric fruit was performed using differential display and Suppression subtractive hybridization. Our analyses resulted in the isolation of 12 differentially expressed cDNAs, which were confirmed by dot-blots and northern blots. Among the sequences identified were sequences homologous to plant aquaporins, adenine nucleotide translocator, immunophilin, legumin-like proteins, deoxyguanosine kinase and omega-3 fatty acid desaturase. Some of these cDNAs correspond to newly isolated genes involved in changes related to the respiratory climacteric, or stress-defense responses. Functional characterization of ripening-associated genes could provide information useful in controlling biochemical pathways that would have an impact on banana quality and shelf life. (C) 2009 Elsevier B.V. All rights reserved.

Role of IL-18 in overt pain-like behaviour in mice

There are evidences that targeting IL-18 might be beneficial to inhibit inflammatory symptoms, including hypernociception (decrease in nociceptive threshold). The mechanism of IL-18 mechanical hypernociception depends on endothelin in rats and mice. However, the role of IL-18 in overt pain-like behaviour remains undetermined. Therefore, we addressed the role of IL-18 in writhing response induced by intraperitoneal (i.p.) injection of phenyl-p-benzoquinone (PBQ) and acetic acid in mice. Firstly, it was detected that PBQ and acetic acid i.p. injection induced a dose-dependent number of writhes in Balb/c mice. Subsequently, it was observed that the PBQ- but not the acetic acid-induced writhes were diminished in IL-18 deficient ((-/-)) mice. Therefore, considering that IFN-gamma, endothelin and prostanoids mediate IL-18-induced mechanical hypernociception, we also investigated the role of these mediators in the same model of writhing response in which IL-18 participates. It was noticed that PBQ-induced writhes were diminished in IFN-gamma(-/-) mice and by the treatment with bosentan (mixed enclothelin ETA/ETB receptor antagonist), BQ 123 (cyclo[DTrp-DAsp-Pro-DVal-Leu], selective enclothelin ETA receptor antagonist), BQ 788 (N-cys-2,6-dimethylpiperidinocarbonyl-L-methylleucyl-D-1 -methoxycarboyl-D-norleucine, selective endothelin ETB receptor antagonist) or indomethacin (cycloxigenase inhibitor). Thus, IL-18, IFN-gamma, endothelin acting on endothelin ETA and ETB receptors, and prostanoids mediate PBQ-induced writhing response in mice. To conclude, these results further advance the understanding of the physiopathology of overt pain-like behaviour, and suggest for the first time a role for IL-18 in writhing response in mice. (C) 2008 Elsevier B.V. All rights reserved.

Antinociceptive activity and toxicology of the lectin from Canavalia boliviana seeds in mice

The aim of the present study was to evaluate the potential antinociceptive and toxicity of Canavalia boliviana lectin (CboL) using different methods in mice. The role of carbohydrate-binding sites was also investigated. CboL given to mice daily for 14 days at doses of 5 mg/kg did not cause any observable toxicity. CboL (1, 5, and 10 mg/kg) administered to mice intravenously inhibited abdominal constrictions induced by acetic acid and the two phases of the formalin test. In the hot plate and tail immersion tests, the same treatment of CboL induced significant increase in the latency period. In the hot plate test, the effect of CboL (5 mg/kg) was reversed by naloxone (1 mg/kg), indicating the involvement of the opioid system. In the open-field and rota-rod tests, the CboL treatment did not alter animals` motor function. These results show that CboL presents antinociceptive effects of both central and peripheral origin, involving the participation of the opioid system via lectin domain.

Stereoselective Analysis of Labetalol in Human Plasma by LC-MS/MS: Application to Pharmacokinetics

Labetalol is clinically available as a mixture of two racemates (four stereoisomers). The stereoisomer (R,R) has as main activity the beta(1)-antagonism and the stereoisomer (S,R) is highly selective for the alpha(1) adrenoceptor and is responsible for most of the alpha-blocker activity. In the present investigation, a method for the analysis of labetalol stereoisomers in human plasma was developed and applied to pharmacokinetic studies. Plasma samples (0.5 ml) were extracted with methyl tert-butyl ether at pH 9.5. The four labetalol stereoisomers were analyzed by LC-MS/MS on a Chirobiotic (R) V column using a mobile phase consisting of methanol, acetic acid, and diethylamine, with a recovery of more than 90% for all four. The quantitation limit was 0.5 ng/ml and linearity was observed at 250 ng/ml plasma for each stereoisomer. Studies of precision and accuracy presented coefficients of variation and percentage inaccuracy of less than 15%, indicating that the method is precise and accurate. The method was applied to the study of the kinetic disposition of labetalol over a period of 12 h after oral administration of a single 100 mg dose to a hypertensive pregnant woman. The clinical study revealed stereoselectivity in the pharmacokinetics of labetalol, with a lower plasma proportion for the active stereoisomers (R,R)-labetalol and (S,R)-labetalol. The stereoselectivity observed after oral administration is due to the hepatic metabolism and the first pass effect, with an AUC((R,R))/AUC((S,S)) ratio of 0.5. Chirality 21:738-744, 2009. (C) 2008 Wiley-Liss, Inc.