Marta M. P. S. Neves

Celiac disease detection using a transglutaminase electrochemical immunosensor fabricated on nanohybrid screen-printed carbon electrodes

Celiac disease is a gluten-induced autoimmune enteropathy characterized by the presence of tissue tranglutaminase (tTG) autoantibodies. A disposable electrochemical immunosensor (EI) for the detection of IgA and IgG type anti-tTG autoantibodies in real patients samples is presented. Screen-printed carbon electrodes (SPCE) nanostructurized with carbon nanotubes and gold nanoparticles were used as the transducer surface. This transducer exhibits the excellent characteristics of carbon-metal nanoparticle hybrid conjugation and led to the amplification of the immunological interaction. The immunosensing strategy consisted of the immobilization of tTG on the nanostructured electrode surface followed by the electrochemical detection of the autoantibodies present in the samples using an alkaline phosphatase (AP) labelled anti-human IgA or IgG antibody. The analytical signal was based on the anodic redissolution of enzymatically generated silver by cyclic voltammetry. The results obtained were corroborated with a commercial ELISA kit indicating that the electrochemical immunosensor is a trustful analytical screening tool.

Celiac disease diagnosis and gluten-free food analytical control

Celiac disease (CD) is an autoimmune enteropathy, characterized by an inappropriate T-cell-mediated immune response to the ingestion of certain dietary cereal proteins in genetically susceptible individuals. This disorder presents environmental, genetic, and immunological components. CD presents a prevalence of up to 1% in populations of European ancestry, yet a high percentage of cases remain underdiagnosed. The diagnosis and treatment should be made early since untreated disease causes growth retardation and atypical symptoms, like infertility or neurological disorders. The diagnostic criteria for CD, which requires endoscopy with small bowel biopsy, have been changing over the last few decades, especially due to the advent of serological tests with higher sensitivity and specificity. The use of serological markers can be very useful to rule out clinical suspicious cases and also to help monitor the patients, after adherence to a gluten-free diet. Since the current treatment consists of a life-long gluten-free diet, which leads to significant clinical and histological improvement, the standardization of an assay to assess in an unequivocal way gluten in gluten-free foodstuff is of major importance.

Synthesis of a Natural Chalcone and Its Prenyl Analogs - Evaluation of Tumor Cell Growth-Inhibitory Activities, and Effects on Cell Cycle and Apoptosis

Six prenyl (=3‐methylbut‐2‐en‐1‐yl) chalcones (=1,3‐diphenylprop‐2‐en‐1‐ones), 2–7, and one natural non‐prenylated chalcone, 1, have been synthesized and evaluated for their in vitro growth‐inhibitory activity against three human tumor cell lines. A pronounced dose‐dependent growth‐inhibitory effect was observed for all prenylated derivatives, except for 7. The chalcone possessing one prenyloxy group at C(2′), i.e., 2, was the most active derivative against the three human tumor cell lines (5.9

An electrochemical deamidated gliadin antibody immunosensor for celiac disease clinical diagnosis

The first electrochemical immunosensor (EI) for the detection of antibodies against deamidated gliadin peptides (DGP) is described here. A disposable nanohybrid screen-printed carbon electrode modified with DGP was employed as the transducers sensing surface. Real serum samples were successfully assayed and the results were corroborated with an ELISA kit. The presented EI is a promising analytical tool for celiac disease diagnosis.

Direct Electroanalytical Determination of Fluvastatin in a Pharmaceutical Dosage Form: Batch and Flow Analysis

Abstract The reduction of luvastatin (FLV) at a hanging mercury-drop electrode (HMDE) was studied by square-wave adsorptive-stripping voltammetry (SWAdSV). FLV can be accumulated and reduced at the electrode, with a maximum peak current intensity at a potential of approximately −1.26 V vs. AgCl/Ag, in an aqueous electrolyte solution of pH 5.25. The method shows linearity between peak current intensity and FLV concentration between 1.0 × 10−8 and 2.7 × 10−6 mol L−1. Limits of detection (LOD) and quantification (LOQ) were found to be 9.9 × 10−9 mol L−1 and 3.3 × 10−8 mol L−1, respectively. Furthermore, FLV oxidation at a glassy carbon electrode surface was used for its hydrodynamic monitoring by amperometric detection in a flow-injection system. The amperometric signal was linear with FLV concentration over the range 1.0 × 10−6 to 1.0 × 10−5 mol L−1, with an LOD of 2.4 × 10−7 mol L−1 and an LOQ of 8.0 × 10−7 mol L−1. A sample rate of 50 injections per hour was achieved. Both methods were validated and showed to be precise and accurate, being satisfactorily applied to the determination of FLV in a commercial pharmaceutical.

Static and hydrodynamic monitoring of citalopram based on its electro-oxidation behavior at a glassy-carbon surface

Abstract The electrooxidative behavior of citalopram (CTL) in aqueous media was studied by cyclic voltammetry (CV) and square-wave voltammetry (SWV) at a glassy-carbon electrode. The electrochemical behaviour of CTL involves two electrons and two protons in the irreversible and diffusion controlled oxidation of the tertiary amine group. The maximum analytical signal was obtained in a phosphate buffer (pH = 8.2). For analytical purposes, an SWV method and a flow-injection analysis (FIA) system with amperometric detection were developed. The optimised SWV method showed a linear range between 1.10 × 10−5–1.20 × 10−4 mol L−1, with a limit of detection (LOD) of 9.5 × 10−6 mol L−1. Using the FIA method, a linear range between 2.00 × 10−6–9.00 × 10−5 mol L−1 and an LOD of 1.9 × 10−6 mol L−1 were obtained. The validation of both methods revealed good performance characteristics confirming applicability for the quantification of CTL in several pharmaceutical products.

Miniaturized analytical instrumentation for electrochemiluminescence assays: a spectrometer and a photodiode-based device

AbstractHerein, a new miniaturized analytical instrumentation for electrochemiluminescence (ECL) assays is presented. A photodiode integrated in an ECL cell combined with a potentiostat/galvanostat, all integrated in a one-piece instrument (μSTAT ECL), was developed. In addition, a complementary micro-spectrometer integrated in a similar ECL cell for luminescence spectra recording is also proposed. Both cells are intended to be used with screen-printed electrodes and all the devices are portable and small sized. Their performance was corroborated with two innovative proofs-of-concept that centered on the luminol transduction chemistry: a first time reported ECL assay based on the enzymatic reaction between an indoxyl substrate and the enzyme alkaline phosphatase, and the electrochemiluminescence resonance energy transfer (ECL-RET) process triggered by the electro-oxidized luminol to the acceptor fluorescein. The photodiode system revealed to be more sensitive than the spectrometer device in collecting the light; however, with the latter, it is possible to discriminate different luminescent species according to their maximum wavelength emission, which is extremely useful for carrying out simple and simultaneous ECL multiplex analyzes. The spectrometer device works as an excellent accessory to couple with the μSTAT ECL instrument, complementing the experiments. Graphical abstractSchematic representation of the ECL-RET: from luminol–H2O2 system to fluorescein, the micro-spectrometer for the light collection and the 3D representation of the ECL-RET reaction.

Potential small-molecule activators of caspase-7 identified using yeast-based caspase-3 and -7 screening assays

Caspases-3 and -7 are at the core of the execution phase of apoptosis. The search for activators of these proteases has therefore deserved particular attention in the field of anticancer drug discovery. Here, a simplified yeast-based screening approach was developed and used to search for activators of caspases-3 and -7, followed by evaluation of the activity of the selected compounds in the human tumor cell lines HL-60 (acute promyelocytic leukemia) and MCF-7 (breast adenocarcinoma). By using the yeast approach, two potential activators of caspase-7, 5,6-dihydroxy-7-prenyloxyflavone (1a) and 3-hydroxy-7-geranyloxyflavone (2a), were identified. Unlike the known caspases-3 and -7 activator, the procaspase activating compound-1 (PAC-1), these flavonoids did not interfere with the caspase-3 activity in yeast. Moreover, flavonoids 1a and 2a processed procaspase-7 to the active caspase-7 both in yeast and in vitro processing assays, and inhibited the growth of HL-60 and MCF-7 human tumor cells with higher potencies than PAC-1, particularly in the absence of caspase-3 (MCF-7 cells). In MCF-7 cells, the flavonoids processed procaspase-7, increased its activity and sensitized these cells to the effects of the cytotoxic drug, etoposide. In conclusion, the developed yeast target-based screening assays led to the identification of potential caspase-7 activators. A proof of concept is therefore provided for the effectiveness of the yeast assays in the discovery of caspase activators. Additionally, the identified compounds may pave the way for a new class of caspase activators with improved anticancer properties.

A non-enzymatic ethanol sensor based on a nanostructured catalytic disposable electrode

Herein, a simple and fast method for the electrocatalytic detection of ethanol using disposable nanostructured screen-printed carbon electrodes (SPCEs) is presented for the first time. Platinum nanoparticles (PtNPs), prepared in the presence of citrate and later purified and dispersed in ultrapure water, were employed in the modification of the SPCEs (SPCE-PtNPs). The synthetized nanoparticles and the catalytic nanostructured solid surface were characterized with transmission electron microscopy (TEM) and with scanning electron microscopy (SEM), respectively. Both systems were also characterized using voltammetric techniques. Finally, the PtNPs modified SPCEs were employed for the electro-oxidation of ethanol in an alkaline medium using portable instrumentation. The electrochemical results revealed that PtNPs can effectively enhance the electron transfer between the analyte of interest and the electrode. The content of ethanol was assayed in real samples (alcoholic beverages) revealing an accurate performance. Additionally, a stability study of the nanostructured surface was carried out. The results obtained corroborate the promising catalytic activity of PtNPs in ethanol detection using disposable, cost-effective and miniaturized sensing devices.

P2.1.24 Multiplexed Electrochemical Immunosensor for Detection of Celiac Disease Serological Markers

Celiac disease is a gluten-induced autoimmune enteropathy characterized by the presence of antibodies against gliadin (AGA) and anti-tissue transglutaminase (anti-tTG) autoantibodies. A disposable electrochemical dual immunosensor for the simultaneous detection of IgA and IgG type AGA and anti-tTG antibodies in real patient’s samples is presented. A screen-printed carbon electrode with two working electrodes nanostructured with a carbon-metal hybrid system worked as the transducer surface. The immunosensing strategy consisted of the immobilization of gliadin and tTG (i.e. CD specific antigens) on the nanostructured electrode surface followed by the electrochemical detection of the human antibodies present in the serum samples assayed. The antigen-antibody interaction was recorded using alkaline phosphatase labelled anti-human antibodies and a mixture of 3-indoxyl phosphate with silver ions (3-IP/Ag+) was used as the substrate. The analytical signal was based on the anodic redissolution of enzymatically generated silver by cyclic voltammetry. The results obtained were corroborated with a commercial ELISA kit indicating that the electrochemical immunosensor is a trustful analytical screening tool.