Diego Centonze

Determination of aflatoxins in cereal flours by solid-phase microextraction coupled with liquid chromatography and post-column photochemical derivatization-fluorescence detection.

A new method for the determination of aflatoxins B(1), B(2), G(1), and G(2) (AFB(1), AFB(2), AFG(1), AFG(2)) in cereal flours based on solid-phase microextraction (SPME) coupled with high performance liquid chromatography with post-column photochemical derivatization and fluorescence detection (SPME-HPLC-PD-FD) has been developed. Aflatoxins were extracted from cereal flour samples by a methanol:phosphate buffer (pH 5.8, I=0.1) (80:20, v/v) solution, followed by a SPME step. Different SPME and HPLC-PD-FD parameters (fiber polarity, temperature, pH, ionic strength, adsorption and desorption time, mobile phase) have been investigated and optimized. This method, which was assessed for the analysis of different cereal flours, showed interesting results in terms of LOD (from 0.035 to 0.2 ng g(-1)), LOQ (from 0.1 to 0.63 ng g(-1), respectively), within and inter-day repeatability (2.27% and 5.38%, respectively) linear ranges (up to 20 ng g(-1) for AFB(1) and AFG(1) and 6 ng g(-1) for AFB(2) and AFG(2)), and total raw extraction efficiency (in the range 55-59% at concentrations in the range 0.3-1 ng g(-1) and 49-52% at concentrations in the range 1-10 ng g(-1)). The results were also compared with the purification step carried out by conventional immunoaffinity columns.

Development of a new analytical method for the determination of sulfites in fresh meats and shrimps by ion-exchange chromatography with conductivity detection

An accurate and reliable analytical method, based on ion chromatography and suppressed conductivity detection, has been developed and validated for the quantitative determination of sulfites in fresh meats and shrimps. The chromatographic separation was accomplished by using an anion-exchange column eluted with sodium carbonate and sodium hydroxide. The optimized step-change elution, followed by column re-equilibration at the initial mobile phase composition, guaranteed a good resolution even toward endogenous interfering peaks, and an excellent retention time repeatability (1.1%, n=6). Good results in terms of sample extract stability, recovery efficiency were achieved with an extraction solvent mixture based on sodium hydroxide, fructose and EDTA. The method validation, performed by an in-house model according to Decision 657/2002/EC and Regulation 882/2004/EC, provided excellent results with respect to linearity (correlation coefficient up to 0.9998), limits of detection and quantification (2.7 and 8.2 mg kg(-1), respectively, expressed as SO(2)), expanded measurement uncertainty (below 10%), recovery values (ranging from 85% to 92%) and repeatability (down to 8%), demonstrating the conformity of the proposed method with the European directives. Finally, by major changes ruggedness studies, the method applicability to the quantitative analysis of cow hamburger, pork and horse sausage, and shrimps was demonstrated.

Multi-residue method for the determination of organochlorine pesticides in fish feed based on a cleanup approach followed by gas chromatography–triple quadrupole tandem mass spectrometry

A multi-residue method for the determination of organochlorine pesticides in fish feed samples was developed and optimized. The method is based on a cleanup step of the extracted fat, carried out by liquid-liquid extraction on diatomaceous earth cartridge with n-hexane/acetonitrile (80/20, v/v) followed by solid phase extraction (SPE) with silica gel-SCX cartridge, before the identification and quantification of the residues by gas chromatography-triple quadrupole tandem spectrometry (GC-MS/MS). Performance characteristics, such as accuracy, precision, linear range, limits of detection (LOD) and quantification (LOQ), for each pesticide were determined. Instrumental LODs ranged from 0.01 to 0.11 microg L(-1), LOQs were in the range of 0.02-0.35 microg L(-1), and calibration curves were linear (r2>0.999) in the whole range of explored concentrations (5-100 microg L(-1)). Repeatability values were in the range of 3-15%, evaluated from the relative standard deviation of six samples spiked at 100 microg kg(-1) of fat, and in compliance with that derived by the Horwitzs equation. No matrix effects or interfering substances were observed in fish feed analyses. The proposed method allowed high recoveries (92-116%) of spiked extracted fat samples at 100 microg kg(-1), and very low LODs (between 0.02 and 0.63 microg kg(-1)) and LOQs (between 0.05 and 2.09 microg kg(-1)) determined in fish feed samples.

Electrochemical immobilisation of enzymes on conducting organic salt electrodes : characterisation of an oxygen independent and interference-free glucose biosensor

Abstract A glucpse biosensor, based on the electrochemical immobilisation of glucose oxidase (GOD) in a poly( o -phenylendiamine) (PPD) film synthesized onto a NMP · TCNQ conducting organic salt (COS) electrode, is described and its performances are evaluated. The electrochemical cycling of the bare conducting organic salt electrode, in a phosphate buffer (pH 7) supporting electrolyte, accumulated a surface layer of TCNQ 0 , which was supposed to be the active mediator for the heterogeneous reoxidation of GOD. Under certain boundary conditions. Chen et al.s model [C.J. Chen, C.C. Liu, R. Savinell, J. Electroanal. Chem., 248 (1993) 317] has been applied to describe the catalysis occurring at our biosensor. Moreover, when the electrolysis is fast, such a model was demonstrated to be equivalent to Albery et al.s mathematics [W.J. Albery, P.N. Bartlett, D.H. Craston, J. Electroanal Chem., 194 (1985) 223], which allows the estimation of the kinetic constants. Such parameters indicated that substrate diffusion through the membrane and unsaturated enzyme kinetics were the rate-limiting processes. COS|PPD|GOD electrodes respond rapidly to glucose, the steady-state current being reached in 10–16 s. Biosensor responses under different operating conditions such as applied potential, presence of oxygen or interferents were investigated. The sensor response, tested in a day-to-day experiment, remained quite stable for 4 days, then decreased to 60% of the initial value on day 5. The shelf lifetime was at least 1 month.

Validation of a confirmatory analytical method for the determination of aflatoxins B1, B2, G1 and G2 in foods and feed materials by HPLC with on-line photochemical derivatization and fluorescence detection

A sensitive and selective analytical method for the simultaneous separation and quantitative determination of aflatoxins B1, B2, G1 and G2 in foodstuffs and materials for feed has been validated. The method is based on high performance liquid chromatography with on-line post-column photochemical derivatization and fluorescence detection. The chromatographic separation of aflatoxins was accomplished using a C18 column eluted with an isocratic mobile phase consisting of water, methanol and acetonitrile. The sample preparation required a simple extraction of aflatoxins with MeOH/H2O (80:20, v/v) and a purification step by immunoaffinity column cleanup. The total analysis time, including sample preparation and chromatographic separation, did not exceed 40 min with a run time of 10 min. The on-line photochemical derivatization ensures better results in terms of simplicity, sensitivity and reproducibility with respect to chemical derivatization techniques, and provides an increase of the peak resolution and an extent of automation in comparison with the electrochemical ones. The procedure for the determination of aflatoxins in food samples and cereals for animal consumption was extensively validated following Regulation (EC) No. 882/2004. Detection limits in wheat bran samples of 0.08 µg kg−1 for AFB1, 0.02 µg kg−1 for AFB2, 0.16 µg kg−1 for AFG1 and 0.04 µg kg−1 for AFG2 were attained. The method allows high recovery with mean values ranging from 72 to 94% and it satisfies the necessary requirements for sensitivity, linearity, selectivity, precision and ruggedness, demonstrating the conformity of the method with provisions of Regulation (EC) No. 401/2006.

Development of a carbon composite electrode made from polyethylene and graphite powder modified with copper(I) oxide

Abstract Solid carbon composite electrodes (CCEs) prepared by copper(I) oxide (Cu 2 O) dispersed in polyethylene and graphite powder have been investigated for use as working electrode in amperometric flow-through detection in alkaline solutions. The effects of matrix composition, electrocatalyst loading, short- and long-term electrode stability, noise current, and sensitivity were assessed using d -mannitol as a model compound. Excellent sensitivity accompanied by a low residual current was observed with a 50% (w/w) Cu 2 O loading level in the carbon paste matrix. An estimate value of the double layer capacitance, evaluated by cyclic voltametry in 0.48 M NaOH, was about 1360 μF/mm 2 . Upon optimizing the sensing electrode in terms of composite electrode formulation, hydroxide ion concentration, and operating potential, the detection limit for d -mannitol in flow injection mode, using a wall-jet flow cell, was 5 μM for 50 μl injections. The bulk Cu 2 O-CCEs exhibit good mechanical stability under forced flow hydrodynamic conditions, and overall offers the advantage of a very low background current. An additional inherent feature of these catalyst-modified electrodes is their renewal capability; the surface can rapidly be renewed by a simple polishing procedure making the amperometric sensor very attractive for routine quantitative measurements.

Voltammetric and XPS investigations of polynuclear ruthenium-containing cyanometallate film electrodes

Abstract Glassy carbon electrodes coated with thin films of mixed valence ruthenium-oxo-species stabilized by cyanoruthenate or cyanoferrate crosslinks were investigated by cyclic voltammetry and X-ray photoelectron spectroscopy (XPS). Electrochemical deposition was accomplished in dilute mineral acid solutions (e.g. 10 mM H 2 SO 4 ) containing millimolar concentrations of RuCl 3 + K 4 Fe(CN) 6 , or RuCl 3 + K 4 Ru(CN) 6 , or OsCl 3 + K 4 Ru(CN) 6 . Film deposition is attributed to growth of compact and conducting lattices with significant oxo- (MOM) and cyano-bridging (M′CNM), in which M represents Ru, or Os, while M′ refers to Ru or Fe. Particular attention was devoted to the polynuclear ruthenium oxide microstructure (RuORu) crosslinked with cyanoruthenate and cyanoferrate, designated as RuORuCN and RuoFeCN respectively. From an electrochemical point of view, the mixed metal RuOFeCN behaves in a manner analogous to the RuORuCN film electrode, and indeed these modified electrodes are capable of catalyzing the oxidation of several substrates in acidic solutions. Depending on the applied potential, two distinct and selective reactive sites are recognizable in the electro-oxidation processes of As(III) and 2-furaldehyde at both RuOFeCN and RuORuCN film electrodes. XPS characterization of these ruthenium-containing cyanometallate films shows clear evidence of the existence of two oxidation states of ruthenium (Ru 3d region), which are respectively attributed to oxo-ruthenium(IV) and dioxo-ruthenium(VI) or Ru(V) in the oxide framework of the inorganic lattice.

Electrocatalysis and amperometric detection of ethanol at ruthenium-based inorganic films with improved response stability

Abstract A glassy carbon electrode coated with a thin film of mixed-oxidation state ruthenium oxide cross-linked with ruthenium cyanide is used for the electrocatalytic oxidation of ethanol in acidic media. This inorganic redox film offers attractive features for application as an electrocatalytic amperometric sensor in batch and flowing solutions. The ruthenium-based modified glassy carbon electrode was assembled in a thin-layer electrochemical cell, and the stability of the film was evaluated. For constant-potential amperometric detection (+ 1.05 V vs. Ag AgCl ) in flow injection analysis, substantial improvements in the response stability of the film were obtained using carrier electrolytes spiked with 5 × 10−6 M of each of the electroplating salts, i.e., RuCl3 and K4Ru(CN)6. Under optimal experimental conditions the peak current response increases linearly with ethanol over the range 0.1–10 mM; linear least squares fit of the data (n = 7) yielded a slope of 0.601 ± 0.003 μA mM−1 (95% confidence limits) with a correlation coefficient of 0.99995.

Comparative analysis of gluten proteins in three durum wheat cultivars by a proteomic approach.

The gluten protein composition and expression level influence dough properties and are cultivar and environment dependent. To broaden the knowledge of the durum wheat gluten proteome, three cultivars were compared in two different growing seasons by a proteomic approach. Cultivar-specific and differentially expressed spots in the two years were identified by mass spectrometry. Significant differences were observed among the cultivars: Ofanto showed the lowest protein spot volumes in the high molecular weight (HMW) and low molecular weight (LMW) <35,000 regions and the highest in the LMW 48,000-35,000 region, Latino the lowest in the LMW 48,000-35,000 region, and Simeto an intermediate expression level in both LMW regions. In the warmer year the up-regulation of HMW glutenins, α-gliadins, and a globulin 3 protein and the down-expression of LMW glutenins and γ-gliadins were observed. Among the cultivars, Simeto showed the highest stability across the environments.

Nickel oxide dispersed in a graphite/poly(vinyl chloride) composite matrix for an electrocatalytic amperometric sensor of alditols in flow-injection analysis

Abstract A novel chemically modified electrode prepared from nickel oxide dispersed in a graphite powder and poly(vinyl chloride) (PVC) composite matrix (NiO-CME) was used for the electrocatalytic oxidation of alditols such as xylitol, sorbitol and mannitol, etc. in alkaline solutions. This bulk-phase modified electrode is well prepared in less than 15 min and its electrode surface is very robust, stable and easily polishable. The detector response was evaluated with regard to flow rate, applied potential and catalyst loading. Amperometric detection in the flow-injection mode at the NiO-CME, using a wall-jet flow cell, showed a linear response up to 50 mM and detection limits of 20 μM, 50 μM and 80 μM for xylitol, dulcitol, and mannitol, respectively. The performance of the sensor was assessed with respect to stability and sensitivity. A signal loss of less than 5% was observed during an operating period of 72 h.