Rifampicin determination in plasma by stir bar-sorptive extraction and liquid chromatography

A sensitive and reproducible stir bar-sorptive extraction and high performance liquid chromatography-UV detection (SBSE/HPLC-UV) method for therapeutic drug monitoring of rifampicin in plasma samples is described and compared with a liquid:liquid extraction (LLE/HPLC-UV) method. This miniaturized method can result in faster analysis, higher sample throughput, lower solvent consumption and less workload per sample while maintaining or even improving sensitivity. Important factors in the optimization of SBSE efficiency such as pH, temperature, extraction time and desorption conditions (solvents, mode magnetic stir, mode ultrasonic stir, time and number of steps) were optimized recoveries ranging from 75 to 80%. Separation was obtained using a reverse phase C(8) column with UV detection (254 nm). The mobile phase consisted of methanol:0.25 N sodium acetate buffer, pH 5.0 (58:42, v/v). The SBSE/HPLC-UV method was linear over a working range of 0.125-50.0 mu g mL(-1). The intra-assay and inter-assay precision and accuracy were studied at three concentrations (1.25, 6.25 and 25.0 mu g mL(-1)). The intra-assay coefficients of variation (CVs) for all compounds were less than 10% and all inter-CVs were less than 10%. Limits of quantification were 0.125 mu g mL(-1). Stability studies showed rifampicin was stable in plasma for 12 h after thawing; the samples were also stable for 24 h after preparation. Based on the figures of merit results, the SBSE/HPLC-UV proved to be adequate to the rifampicin analyses from therapeutic to toxic levels. This method was successfully applied to the analysis of real samples and was as effective as the LLE/HPLC-UV method. (C) 2009 Elsevier B.V. All rights reserved.

Determination of fluoxetine and norfluoxetine enantiomers in human plasma by polypyrrole-coated capillary in-tube solid-phase microextraction coupled with liquid chromatography-fluorescence detection

A sensitive, selective, and reproducible in-tube polypyrrole-coated capillary (PPY) solid-phase microextraction and liquid chromatographic method for fluoxetine and norfluoxetine enantiomers analysis in plasma samples has been developed, validated, and further applied to the analysis of plasma samples from elderly patients undergoing therapy with antidepressants. Important factors in the optimization of in-tube SPME efficiency are discussed, including the sample draw/eject volume, draw/eject cycle number, draw/eject flow-rate, sample pH, and influence of plasma proteins. Separation of the analytes was achieved with a Chiralcel OD-R column and a mobile phase consisting of potassium hexafluorophosphate 7.5 mM and sodium phosphate 0.25 M solution, pH 3.0, and acetonitrile (75:25, v/v) in the isocratic mode, at a flow rate of 1.0 mL/min. Detection was carried out by fluorescence absorbance at Ex/Em 230/290 nm. The multifunctional porous surface structure of the PPY-coated film provided high precision and accuracy for enantiomers. Compared with other commercial capillaries, PPY-coated capillary showed better extraction efficiency for all the analytes. The quantification limits of the proposed method were 10 ng/mL for R- and S-fluoxetine, and 15 ng/mL for R- and S-norfluoxetine, with a coefficient of variation lower than 13%. The response of the method for enantiomers is linear over a dynamic range, from the limit of quantification to 700ng/mL, with correlation coefficients higher than 0.9940. The in-tube SPME/LC method can therefore be successfully used to analyze plasma samples from ageing patients undergoing therapy with fluoxetine. (C) 2009 Elsevier B.V. All rights reserved.

Automated determination of rifampicin in plasma samples by in-tube solid-phase microextraction coupled with liquid chromatography

A sensitive and automated method is described for determination of rifampicin in plasma samples for therapeutic drug monitoring by in-tube solid-phase microextraction coupled with liquid chromatography (in-tube SPME/LC). Important factors in the optimization of in-tube SPME are discussed, such as coating type, sample pH, sample draw/eject volume, number of draw/eject cycles, and draw/eject flow rate. Analyte pre-concentrated in the polyethylene glycol phase was directly transferred to the liquid chromatographic column by percolation of the mobile phase, without carryover. The method was linear over the 0.1-100 mu g/mL range, with a linear coefficient value (r(2)) of 0.998. The inter-assay precision presented coefficient of variation <= 1.7%. The effectiveness and practicability of the proposed method are proven by analysis of plasma samples from ageing patients undergoing therapy with rifampicin. (C) 2011 Elsevier B.V. All rights reserved.

A quantitative coronary angiography-matched comparison between a prospective randomised multicentre cutting balloon angioplasty and bare metal stent trial (REDUCE III) and the Rapamycin-Eluting Stent Evaluation At Rotterdam Cardiology Hospital (RESEARCH) study

Aims: There remains significant concern about the long-term safety of drug-eluting stents (DES). However, bare metal stents (BMS) have been used safely for over two decades. There is therefore a pressing need to explore alternative strategies for reducing restenosis with BMS. This study was designed to examine whether IVUS-guided cutting balloon angioplasty (CBA) with BMS could convey similar restenosis rates to DES. Methods and results: In the randomised REstenosis reDUction by Cutting balloon angioplasty Evaluation (REDUCE III) study, 521 patients were divided into four groups based on device and IVUS use before BMS (IVUS-CBA-BMS: 137 patients; Angio-CBA-BMS: 123; IVUS-BA-BMS: 142; and Angio-BA-BMS: 119). At follow-up, the IVUS-CBA-BMS group had a significantly lower restenosis rate (6.6%) than the other groups (p=0.016). We performed a quantitative coronary angiography (QCA) based matched comparison between an IVUS-guided CBA-BMS strategy (REDUCE III) and a DES strategy (Rapamycin-Eluting-Stent Evaluation At Rotterdam Cardiology. Hospital, the RESEARCH study). We matched the presence of diabetes, vessel size, and lesion severity by QCA. Restenosis (>50% diameter stenosis at follow-up) and target vessel revascularisation (TVR) were examined. QCA-matched comparison resulted in 120-paired lesions. While acute gain was significantly greater in IVUS-CBA-BMS than DES (1.65 +/- 0.41 mm vs. 1.28 +/- 0.57 mm, p=0.001), late loss was significantly less with DES than with IVUS-CBA-BMS (0.03 +/- 0.42 mm vs. 0.80 +/- 0.47 mm, p=0.001). However, no difference was found in restenosis rates (IVUS-CBA-BMS: 6.6% vs. DES: 5.0%, p=0.582) and TVR (6.6% and 6.6%, respectively). Conclusions: An IVUS-guided CBA-BMS strategy yielded restenosis rates similar to those achieved by DES and provided an effective alternative to the use of DES.

Influence of Sensory Cues on the Stimulability for Liquid Sounds in Brazilian Portuguese-Speaking Children

Objective: Stimulability is the ability to produce an adequate sound under specific conditions. This study aimed to describe the stimulability of Brazilian Portuguese-speaking children with and without phonological disorders for the production of liquid sounds with the aid of visual and tactile cues. Patients and Methods: The study sample included 36 children between 5; 0 and 11; 6 years of age, 18 with phonological disorder and 18 without any speech-language disorders. Stimulability was measured for syllable imitation. The stimulability test employed includes 63 syllables with the sounds [1], [(sic)], and [(sic)], as well as seven oral vowels. If the subject was unable to imitate a sound, a visual cue was given. When necessary, a tactile cue was also given. Results: The sound [(sic)] required greater use of sensory cues. Children with phonological disorder needed a greater number of cues. Conclusion: The use of sensory cues seemed to facilitate sound stimulability, making it possible for the children with phonological disorder to accurately produce the sounds modeled. Copyright (C) 2009 S. Karger AG, Basel

Anisotropic Reversible Aggregation of Latex Nanoparticles Suspended in a Lyotropic Nematic Liquid Crystal: Effect of Gradients of Biaxial Order

We studied the anisotropic aggregation of spherical latex particles dispersed in a lyotropic liquid crystal presenting three nematic phases; calamitic, biaxial, and discotic. We observed that in the nematic calamitic phase aggregates of latex particles are formed, which become larger and anisotropic in the vicinity of the transition to the discotic phase, due to a coalescence process. Such aggregates are weakly anisotropic and up to 50 mu m long and tend to align parallel to the director field. At the transition to the discotic phase, the aggregates dissociated and re-formed when the system was brought back to the calamitic phase. This shows that the aggregation is due to attractive and repulsive forces generated by the particular structure of the nematic phase. The surface-induced positional order was investigated by surface force apparatus experiments with the lyotropic system confined between mica surfaces, revealing the existence of a presmectic wetting layer around the surfaces and oscillating forces of increasing amplitude as the confinement thickness was decreased. We discuss the possible mechanisms responsible for the reversible aggregation of latex particles, and we propose that capillary condensation of the N(C) phase, induced by the confinement between the particles, could reduce or remove the gradient of order parameter, driving the transition of aggregates from solidlike to liquidlike and gaslike.

Enantioselective Determination of Mexiletine and Its Metabolites p-Hydroxymexiletine and Hydroxymethylmexiletine in Rat Plasma by Normal-Phase Liquid Chromatography-Tandem Mass Spectrometry: Application to Pharmacokinetics

Mexiletine (MEX), hydroxymethylmexiletine (HMM) and P-hydroxy-mexiletine (PHM) were analyzed in rat plasma by LC-MS/MS. The plasma samples were prepared by liquid-liquid extraction using methyl-tert-butyl ether as extracting solvent. MEX, HMM, and PHM enantiomers were resolved on a Chiralpak (R) AD column. Validation of the method showed a relative standard deviation (precision) and relative errors (accuracy) of less than 15% for all analytes studied. Quantification limits were 0.5 ng ml(-1) for the MEX and 0.2 ng ml(-1) for the HMM and PHM enantiomers. The validated method was successfully applied to quantify the enantiomers of MEX and its metabolites in plasma samples of rats (n = 6) treated with a single oral dose of racemic MEX. Chirality 21:648-656, 2009. (C) 2008 Wiley-Liss, Inc.

Lsa63, a newly identified surface protein of Leptospira interrogans binds laminin and collagen IV

Leptospira interrogans is the etiological agent of leptospirosis, a zoonotic disease that affects populations worldwide. We have identified in proteomic studies a protein that is encoded by the gene LIC10314 and expressed in virulent strain of L. interrogans serovar Pomona. This protein was predicted to be surface exposed by PSORT program and contains a p83/100 domain identified by BLAST analysis that is conserved in protein antigens of several strains of Borrelia and Treponema spp. The proteins containing this domain have been claimed antigen candidates for serodiagnosis of Lyme borreliosis. Thus, we have cloned the LIC10314 and expressed the protein in Escherichia coli BL21-SI strain by using the expression vector pAE. The recombinant protein tagged with N-terminal hexahistidine was purified by metal-charged chromatography and characterized by circular dichroism spectroscopy. This protein is conserved among several species of pathogenic Leptospira and absent in the saprophytic strain L. biflexa. We confirm by liquid-phase immunofluorescence assays with living organisms that this protein is most likely a new surface leptospiral protein. The ability of the protein to mediate attachment to ECM components was evaluated by binding assays. The leptospiral protein encoded by LIC10314, named Lsa63 (Leptospiral surface adhesin of 63 kDa), binds strongly to laminin and collagen IV in a dose-dependent and saturable fashion. In addition, Lsa63 is probably expressed during infection since it was recognized by antibodies of serum samples of confirmed-leptospirosis patients in convalescent phase of the disease. Altogether, the data suggests that this novel identified surface protein may be involved in leptospiral pathogenesis. (C) 2009 The British Infection Society. Published by Elsevier Ltd. All rights reserved.

A newly identified protein of Leptospira interrogans mediates binding to laminin

Pathogenic Leptospira is the aetiological agent of leptospirosis, a life-threatening disease that affects populations worldwide. The search for novel antigens that could be relevant in host-pathogen interactions is being pursued. These antigens have the potential to elicit several activities, including adhesion. This study focused on a hypothetical predicted lipoprotein of Leptospira, encoded by the gene LIC12895, thought to mediate attachment to extracellular matrix (ECM) components. The gene was cloned and expressed in Escherichia coli BL21 Star (DE3)pLys by using the expression vector pAE. The recombinant protein tagged with N-terminal hexahistidine was purified by metal-charged chromatography and characterized by circular dichroism spectroscopy. The capacity of the protein to mediate attachment to ECM components was evaluated by binding assays. The leptospiral protein encoded by LIC12895, named Lsa27 (leptospiral surface adhesin, 27 kDa), bound strongly to laminin in a dose-dependent and saturable fashion. Moreover, Lsa27 was recognized by antibodies from serum samples of confirmed leptospirosis specimens in both the initial and the convalescent phases of the disease. Lsa27 is most likely a surface protein of Leptospira as revealed in liquid-phase immunofluorescence assays with living organisms. Taken together, these data indicate that this newly identified membrane protein is expressed during natural infection and may play a role in mediating adhesion of L. interrogans to its host.

EFFECT OF METAL PRIMERS ON MICROTENSILE BOND STRENGTH BETWEEN ZIRCONIA AND RESIN CEMENTS

Statement of problem. There are no established clinical procedures for bonding zirconia to tooth structure using resin cements. Purpose. The purpose of this study was to evaluate the influence of metal primers, resin cements, and aging on bonding to zirconia. Material and methods. Zirconia was treated with commercial primers developed for bonding to metal alloys (Metaltite, Metal Primer II, Alloy Primer or Totalbond). Non-primed specimens were considered as controls. One-hundred disk-shaped specimens (19 x 4 mm) were cemented to composite resin substrates using Panavia or RelyX Unicem (n=5). Microtensile bond strength specimens were tested after 48 hours and 5 months (150 days), and failure modes were classified as type 1 (between ceramic/cement), 2 (between composite resin/cement) or 3 (mixed). Data were analyzed by 3-way ANOVA and Multiple Comparison Tukey test (alpha=.05). Results. The interactions primer/luting system (P=.016) and luting system/storage time (P=.004) were statistically significant. The use of Alloy Primer significantly improved the bond strength of RelyX Unicem (P<.001), while for Panavia, none of the primers increased the bond strength compared to the control group. At 48 hours, Panavia had statistically higher bond strength (P=.004) than Unicem (13.9 +/- 4.4MPa and 10.2 +/- 6.6MPa, respectively). However, both luting systems presented decreasing, statistically similar; values after aging (Panavia: 3.6 +/- 2.2MPa; Unicem: 6.1 +/- 5.3MPa). At 48 hours, Alloy Primer/Unicem had the lowest incidence of type 1 failure (8%). After aging, all the groups showed a predominance of type 1 failures. Conclusions. The use of Alloy Primer improved bond strength between RelyX Unicem and zirconia. Though the initial values obtained with Panavia were significantly higher than RelyX Unicem, after aging, both luting agents presented statistically similar performances. (J Prosthet Dent 2011;105:296-303)

The Effect of Framework Design on Fracture Resistance of Metal-Ceramic Implant-Supported Single Crowns

This study evaluated the effect of framework design on the fracture resistance of metal-ceramic implant-supported crowns. Screw-retained molar crowns with a screw access hole composed of metal or porcelain were compared to a cement-retained crown (control). For each group (n = 10), five crowns were subjected to dynamic loading (1,200,000 x 100 N x 2 Hz at 37 degrees C). Afterward, all specimens were loaded to failure using a universal testing machine. Significant differences could be established between the cement-and screw-retained groups (P <= .05), but no difference was found between the screw-retained groups and the specimens subjected to dynamic loading. Occlusal discontinuity of screw-retained crowns affects their resistance, and the metallic support on the screw access hole did not reinforce the crowns. Int J Prosthodont 2010;23:350-352.

IN VITRO FRACTURE RESISTANCE OF GLASS-FIBER AND CAST METAL POSTS WITH DIFFERENT LENGTHS

Statement of problem. Dental fractures can occur in endodontically treated teeth restored with posts. Purpose. The purpose of this study was to evaluate the in vitro fracture resistance of roots with glass-fiber and metal posts of different lengths. Material and methods. Sixty endodontically treated maxillary canines were embedded in acrylic resin, except for 4 mm of the cervical area, after removing the clinical crowns. The post spaces were opened with a cylindrical bur at low speed attached to a surveyor, resulting in preparations with lengths of 6 mm (group 6 mm), 8 mm (group 8 mm), or 10 mm (group 10 mm). Each group was divided into 2 subgroups according to the post material: cast post and core or glass-fiber post (n=30). The posts were luted with dual-polymerizing resin cement (Panavia F). Cast posts and cores of Co-Cr (Resilient Plus) crowns were made and cemented with zinc phosphate. Specimens were subjected to increasing compressive load (N) until fracture. Data were analyzed with 2-way ANOVA and the Tukey-Kramer test (alpha=.05). Results. The ANOVA analysis indicated significant differences (P<.05) among the groups, and the Tukey test revealed no significant difference among the metal posts of 6-mm length (26.5 N +/- 13.4), 8-mm length (25.2 N +/- 13.9), and 10-mm length (17.1 N +/- 5.2). Also, in the glass-fiber post group, there was no significant difference when posts of 8-mm length (13.4 N +/- 11.0) were compared with the 6-mm (6.9 N +/- 4.6) and 10-mm (31.7 N +/- 13.1) groups. The 10-mm-long post displayed superior fracture resistance, and the 6-mm-long post showed significantly lower mean values (P<.001). Conclusions. Within the limitations of this study, it was concluded that the glass-fiber post represents a viable alternative to the cast metal post, increasing the resistance to fracture of endodontically treated canines. (J Prosthet Dent 2009;101:183-188)