Microcarrier-based fluorescent yeast estrogen screen assay for fast determination of endocrine disrupting compounds.

The presence of endocrine-disrupting compounds (EDCs) in water poses a significant threat to human and animal health, as recognized by regulatory agencies throughout the world. The Yeast Estrogen Screen (YES) assay is an excellent method to evaluate the presence of these compounds in water due to its simplicity and capacity to assess the bioaccessible forms/fractions of these compounds. In the presence of a compound with estrogenic activity, Saccharomyces cerevisiae cells, containing a lacZ reporter gene encoding the enzyme β-galactosidase, are induced, the enzyme is synthesised, and released to the extracellular medium.

Efficacy, safety, and tolerability of adjunctive eslicarbazepine acetate in patients with focal to bilateral tonic-clonic seizures.

Objective: To report the efficacy, safety, and tolerability of adjunctive eslicarbazepine acetate (ESL) treatment in reducing focal to bilateral tonic-clonic seizures (FBTCS).

Methods: Data were pooled from 3 randomized clinical trials (RCTs) of adjunctive ESL in patients with focal seizures. Patients treated with 800 or 1200 mg/day ESL and who experienced ≥ 1 FBTCS during baseline were included.

Antiepileptogenesis after stroke-trials and tribulations: Methodological challenges and recruitment results of a Phase II study with eslicarbazepine acetate.

There is currently no evidence to support the use of antiseizure medications to prevent unprovoked seizures following stroke. Experimental animal models suggested a potential antiepileptogenic effect for eslicarbazepine acetate (ESL), and a Phase II, multicenter, randomized, double-blind, placebo-controlled study was designed to test this hypothesis and assess whether ESL treatment for 1 month can prevent unprovoked seizures following stroke. We outline the design and status of this antiepileptogenesis study, and discuss the challenges encountered in its execution to date.

Safety of Eslicarbazepine Acetate in Elderly Versus Non-Elderly Patients with Focal Seizures: From Pooled Data of Clinical Studies to 8 Years of Post-Marketing Experience.

Introduction: The prevalence of epilepsy increases in elderly patients aged > 65 years, and treatment is challenging because clinical data are limited.

Objective: Our objective was to evaluate the safety of eslicarbazepine acetate (ESL) in patients aged ≥ 65 years versus non-elderly patients with focal seizures.

Methods: The safety data of seven phase II and III, double-blind, open-label, randomized clinical studies of ESL in adults were pooled.

Assessment of immunoglobulin capture in immobilized protein A through automatic bead injection.

The repeatable immobilization of molecular recognition elements onto particle surfaces has a strong impact on the outcomes of affinity-based assays. In this work, an automatic method for the immobilization of immunoglobulin G (IgG) onto protein A-Sepharose microbeads was established through the flow programming features of the portable lab-on-valve platform using micro-bead injection spectroscopy. The reproducible packing of protein A-microbeads between two optic fibers was feasible, allowing on-column probing of IgG retention.

Eslicarbazepine acetate exposure in pregnant women with epilepsy.

Purpose: Epilepsy is a common neurologic disorder requiring continued treatment during pregnancy. Treatment with antiepileptic drugs (AEDs) is needed for seizure control, but the risk of adverse events has to be minimized for both mother and foetus. Available data on pregnancy and foetal/postnatal outcomes following eslicarbazepine acetate (ESL) exposure via parent is herein presented for the first time.

Micro-bead injection spectroscopy for label-free automated determination of immunoglobulin G in human serum.

Immunoglobulin G (IgG) represents the major fraction of antibodies in healthy adult human serum, and deviations from physiological levels are a generic marker of disease corresponding to different pathologies. Therefore, screening methods for IgG evaluation are a valuable aid to diagnostics. The present work proposes a rapid, automatic, and miniaturized method based on UV-vis micro-bead injection spectroscopy (μ-BIS) for the real-time determination of human serum IgG with label-free detection.

Safety Profile of Eslicarbazepine Acetate as Add-On Therapy in Adults with Refractory Focal-Onset Seizures: From Clinical Studies to 6 Years of Post-Marketing Experience.

Introduction: Eslicarbazepine acetate was first approved in the European Union in 2009 as adjunctive therapy in adults with partial-onset seizures with or without secondary generalization.

Objective: The objective of this study was to review the safety profile of eslicarbazepine acetate analyzing the data from several clinical studies to 6 years of post-marketing surveillance.

Methods: We used a post-hoc pooled safety analysis of four phase III, double-blind, randomized, placebo-controlled studies (BIA-2093-301, -302, -303, -304) of eslicarbazepine acetate as add-on therapy in adults.

Dynamic flow-through approach to evaluate readily bioaccessible antioxidants in solid food samples.

Release of bioactive compounds from food matrices is regarded as the first step towards their human bioavailability. The objective of this work was the implementation of an affordable and robust flow-through device for expedient dynamic leaching experiments aiming at the assessment of readily bioaccessible antioxidant compounds in solid food commodities. A simple configuration is proposed using commercially available devices containing regenerated cellulose filters placed in polypropylene holders to entrap the solid sample, featuring a disposable, single use extraction chamber.

Evaluation of the joint effect of the incorporation of mechanically deboned meat and grape extract on the formulation of chicken nuggets.

The proximate composition, instrumental and perceived appearance of chicken nuggets formulated with varying contents of mechanically deboned chicken meat and varying concentrations of grape pomace extract were evaluated, with the choice of formulations following a central composite design. Significant differences (P < 0.05) in fat content were mainly associated to the extent of mechanically deboned chicken meat incorporation.

Do cinnamylideneacetophenones have antioxidant properties and a protective effect toward the oxidation of phosphatidylcholines?

Cinnamylideneacetophenones (CA) are an important group of α,β,γ,δ-diunsaturated ketones that have been widely used in a variety of synthetic transformations. Biological studies concerning these compounds are scarce and refer mainly to antiviral and antibacterial evaluations. Curcumin (CR), a natural polyphenol, is a yellow pigment extracted from the plant Curcuma longa, which is one of the major spices used in the Indian culinary.

Programmable flow system for automation of oxygen radical absorbance capacity assay using pyrogallol red for estimation of antioxidant reactivity.

An automated oxygen radical absorbance capacity (ORAC) method based on programmable flow injection analysis was developed for the assessment of antioxidant reactivity. The method relies on real time spectrophotometric monitoring (540 nm) of pyrogallol red (PGR) bleaching mediated by peroxyl radicals in the presence of antioxidant compounds within the first minute of reaction, providing information about their initial reactivity against this type of radicals. The ORAC-PGR assay under programmable flow format affords a strict control of reaction conditions namely reagent mixing, temperature and reaction timing, which are critical parameters for in situ generation of peroxyl radical from 2,2'-azobis(2-amidinopropane) dihydrochloride (AAPH).

On-line automated evaluation of lipid nanoparticles transdermal permeation using Franz diffusion cell and low-pressure chromatography.

A low-pressure liquid chromatography system for the on-line quantification of caffeine loaded into lipid nanoparticles that permeates pig skin was developed. The apparatus includes a Franz diffusion cell with computer-controlled sampling that allows collection of acceptor solution with automatic compensation for sample withdrawing, and a C-18 reversed-phase monolithic column integrated in a typical Flow Injection Analysis (FIA) set-up where separation between caffeine and other matrix elements is performed before spectrophotometric quantification at 273 nm. Several parameters regarding chromatographic analysis (propulsion element, column length, mobile phase composition, and flow rate) were studied along with the establishment of the sampling procedure.

Rapid assessment of bioactive phenolics and methylxanthines in spent coffee grounds by FT-NIR spectroscopy.

Spent coffee grounds (SCGs) are a great source of bioactive compounds with interest to pharmaceutical and cosmetic industries. Phenolics and methylxanthines are the main health related compounds present in SCG samples. Content estimation of these compounds in SCGs is of upmost importance in what concerns their profitable use by waste recovery industries.

Myoglobin microplate assay to evaluate prevention of protein peroxidation.

The current therapeutic strategies are based on the design of multifunctional drug candidates able to interact with various disease related targets. Drugs that have the ability to scavenge reactive oxygen species (ROS), beyond their main therapeutic action, may prevent the oxidative damage of biomolecules. Therefore, analytical approaches that monitor in a continuous mode the ability of drugs to counteract peroxidation of physiologically relevant biotargets are required.

Automatic flow injection analysis (FIA) determination of total reducing capacity in serum and urine samples.

Automation of total antioxidant capacity assessment can substantially increase the determination throughput, allowing large scale studies and screening experiments. Total reducing capacity evaluation can be implemented under different chemistries, including the CUPRAC-Cupric Ion Reducing Antioxidant Capacity -assay. This assay is based on reduction of Cu(II)-neocuproine complex to highly colored Cu(I)-neocuproine complex by reducing (antioxidant) components of biological samples.

Assessing oral bioaccessibility of trace elements in soils under worst-case scenarios by automated in-line dynamic extraction as a front end to inductively coupled plasma atomic emission spectrometry.

A novel biomimetic extraction procedure that allows for the in-line handing of ≥400 mg solid substrates is herein proposed for automatic ascertainment of trace element (TE) bioaccessibility in soils under worst-case conditions as per recommendations of ISO norms. A unified bioaccessibility/BARGE method (UBM)-like physiological-based extraction test is evaluated for the first time in a dynamic format for accurate assessment of in-vitro bioaccessibility of Cr, Cu, Ni, Pb and Zn in forest and residential-garden soils by on-line coupling of a hybrid flow set-up to inductively coupled plasma atomic emission spectrometry. Three biologically relevant operational extraction modes mimicking: (i) gastric juice extraction alone; (ii) saliva and gastric juice composite in unidirectional flow extraction format and (iii) saliva and gastric juice composite in a recirculation mode were thoroughly investigated.

Insights on antioxidant assays for biological samples based on the reduction of copper complexes-the importance of analytical conditions.

Total antioxidant capacity assays are recognized as instrumental to establish antioxidant status of biological samples, however the varying experimental conditions result in conclusions that may not be transposable to other settings. After selection of the complexing agent, reagent addition order, buffer type and concentration, copper reducing assays were adapted to a high-throughput scheme and validated using model biological antioxidant compounds of ascorbic acid, Trolox (a soluble analogue of vitamin E), uric acid and glutathione. A critical comparison was made based on real samples including NIST-909c human serum certified sample, and five study samples.

Automated microdialysis-based system for in situ microsampling and investigation of lead bioavailability in terrestrial environments under physiologically based extraction conditions.

In situ automatic microdialysis sampling under batch-flow conditions is herein proposed for the first time for expedient assessment of the kinetics of lead bioaccessibility/bioavailability in contaminated and agricultural soils exploiting the harmonized physiologically based extraction test (UBM). Capitalized upon a concentric microdialysis probe immersed in synthetic gut fluids, the miniaturized flow system is harnessed for continuous monitoring of lead transfer across the permselective microdialysis membrane to mimic the diffusive transport of metal species through the epithelium of the stomach and of the small intestine. Besides, the addition of the UBM gastrointestinal fluid surrogates at a specified time frame is fully mechanized.

Rapid assessment of endpoint antioxidant capacity of red wines through microchemical methods using a kinetic matching approach.

Antioxidant capacity of food samples is usually assessed by different analytical methods, however the results attained even for the same method are strongly dependent on the selected reaction time and also on the standard compound used. To tackle this problem, we propose here a kinetic matching approach, associated to the conversion of results into equivalents of a common standard compound, as a universal way for expression of results. The methodology proposed was applied to methods based on different chemistries (Folin-Ciocalteu (F-C), CUPRAC, DPPH(•) and ABTS(•+) assays) and red wines (n=40) were chosen as a model of complex food sample.

High-throughput total cupric ion reducing antioxidant capacity of biological samples determined using flow injection analysis and microplate-based methods.

High-throughput cupric ion reducing antioxidant capacity (CUPRAC) methods were developed for assessment of total antioxidant capacity (TAC) in urine and serum, based on reduction of Cu(II)-neocuproine complex to highly colored Cu(I)-neocuproine complex, measured spectrophotometrically at 450 nm. The reaction time was significantly reduced from 30 to 4 min by application of a calibration compound (uric acid) with kinetic behavior similar to that shown by urine samples. The method was implemented in a microformat (96 well plates) and also in an automatic fashion (flow injection analysis, FIA).

Determination of the scavenging capacity against reactive nitrogen species by automatic flow injection-based methodologies.

Automatic flow-based systems have been applied to assay scavenging capacity against reactive oxygen and nitrogen species, providing analytical tools which can cope with different types and large number of samples. In the present chapter, a flow injection analysis procedure is described for the assessment of peroxynitrite scavenging. A sequential injection analysis procedure is also described for determining the scavenging capacity against the nitric oxide radical.

Rapid microplate high-throughput methodology for assessment of Folin-Ciocalteu reducing capacity.

In the present work, a rapid and high-throughput Folin-Ciocalteu (F-C) reducing capacity assay adapted to routine/screening analysis was developed. In order to attain a fast F-C reducing kinetic reaction, the reaction conditions of the classical time-consuming F-C assay were modified and the influence of alkali and F-C reagent concentration was evaluated using gallic acid as standard. The proposed method was performed in a 96-well microplate format and it was applied to several phenolic compounds and food products (wines, beers, infusions and juices) providing F-C reducing capacity results after 3 min of reaction similar to those obtained by the time-consuming (120 min) conventional method.

High-throughput microplate assay for the determination of drug partition coefficients.

Partition coefficients (K(p)) of drugs between the phospholipid bilayer and the aqueous phase provide useful information in quantitative structure-activity relationship studies. Hexadecylphosphocholine (HePC) micelles, composed of a zwitterionic hydrophilic surface and a hydrophobic core, mimic the biomembranes and have several advantages over other lipid structures to assess K(p) values. Their preparation is easy, fast and avoids the use of toxic organic solvents, and the output has fewer spectroscopic interferences.

Fully automatic flow method for the determination of scavenging capacity against nitric oxide radicals.

In the present work, a fluorimetric automatic method based on multisyringe flow injection analysis (MSFIA) was developed for in vitro evaluation of scavenging capacity against nitric oxide (NO) using 4,5-diaminofluorescein (DAF-2) as an NO-selective fluorogenic probe. The MSFIA manifold was assembled to perform the in-line generation of NO and the competitive reaction of putative scavenger molecules and DAF-2 with NO at conditions close to those found in vivo regarding temperature (37 degrees C), pH (7.4), and concentration of NO (less than 1 microM).