Talanta (v.82, #2)

Thermospray generation directly into a flame furnace—An alternative to improve the detection power in atomic absorption spectrometry by Marcos Almeida Bezerra; Valfredo Azevedo Lemos; Jerusa Simone Garcia; Douglas Gonçalves da Silva; André Souza Araújo; Marco Aurélio Zezzi Arruda (437-443).
Recent developments and applications in the production of thermosprays directly into flame furnaces to improve the analytical sensitivity in atomic absorption spectrometry are reviewed in this manuscript. Principles, characteristics, instrumentation, and applications of this analytical technique for trace elements determination in several matrices are discussed. The use of preconcentration procedures to allow low detection limits for ultra-trace levels using TS-FF-AAS is presented and current perspectives and future trends of this technique are also discussed.
Keywords: TS-FF-AAS; Thermospray; Flame furnace; Volatile elements; Trace analysis; Preconcentration;

Ninhydrin reaction on thiol-reactive solid and its potential for the quantitation of d-penicillamine by Theerasak Rojanarata; Praneet Opanasopit; Tanasait Ngawhirunpat; Choedchai Saehuan (444-449).
While aminothiols produce weak purple-colored reactions with ninhydrin, we demonstrate for the first time that this color could be intensely developed. Using a d-penicillamine paradigm, adsorption of this compound via a disulfide bond onto thiol-reactive solid prior to ninhydrin reaction allowed spectrophotometrical monitoring of the supernatant at 570 nm. Compared with off-solid method, this approach expanded the linear concentration range to 50–600 μg mL−1 and enhanced the sensitivity so that d-penicillamine with the concentrations of less than 100 μg mL−1 could be accurately quantitated by using a second-order polynomial calibration curve. Additionally, this assay was unaffected by disulfide adduct interference, highlighting its potential for the analysis of d-penicillamine as well as other aminothiols.
Keywords: Ninhydrin; Thiol-reactive solid; Penicillamine; Aminothiol;

Synthesis and characteristic of the Fe3O4@SiO2@Eu(DBM)3·2H2O/SiO2 luminomagnetic microspheres with core-shell structure by Ping Lu; Ji-Lin Zhang; Yan-Lin Liu; De-Hui Sun; Gui-Xia Liu; Guang-Yan Hong; Jia-Zuan Ni (450-457).
The Fe3O4@SiO2@Eu(DBM)3·2H2O/SiO2 microsphere composed of Fe3O4 particulate as a magnetic core and silica doped with fluorescent europium complex as an outer shell was fabricated. The obtained composite microspheres with hypersensitive red fluorescence, strong magnetic response, and good water-dispersibility have potential biomedical applications in bioimaging.The core-shell structured luminomagnetic microsphere composed of a Fe3O4 magnetic core and a continuous SiO2 nanoshell doped with Eu(DBM)3·2H2O fluorescent molecules was fabricated by a modified Stöber method combined with a layer-by-layer assembly technique. X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FTIR), field emission scanning electron microscopy (FE-SEM), transmission electron microscopy (TEM), confocal microscopy, photoluminescence (PL), and superconducting quantum interface device (SQUID) were employed to characterize the Fe3O4@SiO2@Eu(DBM)3·2H2O/SiO2 microspheres. The experimental results show that the microshpere has a typical diameter of ca. 500 nm consisting of the magnetic core with about 340 nm in diameter and silica shell doped with europium complex with an average thickness of about 80 nm. It possesses magnetism with a saturation magnetization of 25.84 emu/g and negligible coercivity and remanence at room temperature and exhibits strong red emission peak originating from electric-dipole transition 5D0  →  7F2 (611 nm) of Eu3+ ions. The luminomagnetic microspheres can be uptaken by HeLa cells and there is no adverse cell reaction. These results suggest that the luminomagnetic microspheres with magnetic resonance response and fluorescence probe property may be useful in biomedical imaging and diagnostic applications.
Keywords: Fluorescence; Magnetic; Microsphere; Cell imaging; Eu(III) complex; Magnetite;

Enhanced gas sensing by assembling Pd nanoparticles onto the surface of SnO2 nanowires by Hui Li; Jiaqiang Xu; Yongheng Zhu; Xuedong Chen; Qun Xiang (458-463).
SnO2 nanowires with an average 0.6 μm in length and about 25 nm in diameter were prepared by a hydrothermal method. The sensors were fabricated using SnO2 nanowires assembled with Pd nanocrystals. The sensing properties of the sensors such as selectivity, response–recovery time and stability were tested at 290 °C. After assembling Pd nanocrystals onto the surface of SnO2 nanowires, the gas sensing properties of the sensors toward H2S were improved. The sensors based on Pd nanoparticle@SnO2 nanowires exhibit high stability owing to stable single crystal structure. The mechanism of promoting sensing properties with Pd nanoparticles is discussed.
Keywords: SnO2 nanowires; Pd nanoparticles; Gas sensing; Self-assembly;

A novel stir bar coated with molecularly imprinted polymer (MIP) as selective extraction phase for sorptive extraction of triazine herbicides was developed. The stir bar was prepared by chemically bonding the MIP to the glass bar to improve its stability. A homogeneous and porous structure was observed on the stir bar surface. Extraction performance shows that the MIP-coated stir bar has stronger affinity to the template molecule terbuthylazine as compared with that of the reference stir bar without addition of template. Owning to the shape and structural compatibility, the obtained stir bar also demonstrated specific selectivity to the structural related-compounds of nine triazines, and thus can be applied to simultaneous determination of these compounds from complex samples coupled with high performance liquid chromatography. Four complex samples with different matrix, including rice, apple, lettuce and soil were used to evaluate this proposed method. The limits of detection obtained are in the range of 0.04–0.12 μg L−1, and the recoveries for the spiked rice, apple, lettuce and soil samples were 80.8–107.7%, 80.6–107.8%, 72.0–109.8% and 89.0–114.8% with RSD from 1.2 to 7.9%, respectively. Moreover, this MIP-coated stir bar was firm, durable and can be prepared simply and reproducibly. The developed coating method would be useful to prepare a range of selective stir bars in order to extend the applicability of stir bar sorptive extraction (SBSE) in complex sample analysis.
Keywords: Stir bar sorptive extraction; Molecularly imprinted; Complex sample; Triazines;

A simple in situ solvent formation microextraction methodology based on the application of ionic liquid (IL) as an extractant solvent and sodium hexafluorophosphate (NaPF6) as an ion-pairing agent was proposed for the preconcentration of trace levels of cadmium. In this method cadmium was complexed with O,O-diethyldithiophosphate (DDTP) and extracted into an ionic liquid phase. After phase separation, the enriched analyte in the final solution is determined by flame atomic absorption spectrometry (FAAS). ISFME is a simple and rapid method for extraction and preconcentration of metal ions from sample solutions containing a high concentration of salt. Some effective factors that influence the microextraction efficiency were investigated and optimized. Under the optimum experimental conditions, the limit of detection (3 s) and the enhancement factor were 0.07 μg L−1 and 78, respectively. The relative standard deviation (R.S.D.) was obtained 2.42%. The accuracy of the method was confirmed by analyzing certified reference materials for trace elements in seawater (GBW (E) 080040 seawater). The proposed method was successfully applied for the determination of cadmium in water samples and food grade salts.
Keywords: In situ solvent formation microextraction; Ionic liquid; Cadmium; Flame atomic absorption spectrometry; Saline samples;

Massively parallel genomic DNA fragments display on chip plays a key role in the new generation DNA sequencing. Here, we developed a new technology to display the parallel genomic DNA fragment massively based on two-step reaction with Ф29 DNA polymerase. The genomic DNA fragments were firstly amplified by rolling-circle amplification (RCA) reaction in liquid phase, and then amplified further on the chip by the strand displacement of Ф29 DNA polymerase. In our experiments, through DNA colonies produced by two-step amplification reaction T7 genomic DNA fragments are displayed massively and parallely on the chip, which has been verified through hybridizing the probe labeled with fluorescence or extension reaction with fluorescent-dNTP. The significant difference of fluourescence signals between background and displayed DNA fragments could be obtained. Our results show that the method has good reproducibility in experiments, which may be hopeful to serve the high-throughput sequencing.
Keywords: RCA; Strand displacement amplification; DNA colony;

A sensitive choline biosensor with supramolecular architecture by Zhanxia Zhang; Juan Wang; Xiaolei Wang; Yong Wang; Xiurong Yang (483-487).
A sensitive biosensor with supramolecular architecture was designed and implemented here to detect choline. Choline oxidase and horseradish peroxidase were assembled onto the polymer of thiolated β-cyclodextrin and platinum nanoparticles modified gold electrode through 1-adamantane carboxylic acid coupling. Square wave voltammetry showed that the reduction currents at 0.38 mV had a linear relationship with the logarithms of choline concentrations in the range of 10−9–10−2  M, and the detection limit was down to 0.1 nM. Such biosensor also exhibited excellent selectivity, reproducibility and stability.
Keywords: Choline biosensor; Supramolecular complex; β-Cyclodextrin; Platinum nanoparticles; Square wave voltammetry;

The silica monolith with ionizable silanol groups and large surface area was found able to function as an offline cation exchange solid phase extraction (SPE) cartridge for extracting polar analytes. The prepared cartridge was housed in a 2-mL syringe fixed over a SPE vacuum manifold. The unique property of this silica monolithic cartridge was demonstrated by extracting epinephrine, normetanephrine and metanephrine from urine samples. These analytes were chosen as model compounds for testing because of their high hydrophilicity, and being candidates monitored for clinical diagnosis. The extracted analytes, after concentration and reconstitution were then quantitated by high-performance liquid chromatography coupled to mass spectrometer (HPLC/ESI/MS). Multiple reactions monitoring was carried out with transitions: 184 → 107, 184 → 134 and 198 → 148 for analyzing epinephrine, normetanephrine and metanephrine, respectively. The limit of detection was 3 ng/mL for metanephrine and 5 ng/mL for normetanephrine and epinephrine. The relative standard deviations of measurements ranged from 2 to 10%. The sorbent offered good linearity with coefficient of determination (r 2) > 0.99, over a concentration range of 20–200 ng/mL. The relative recoveries ranged from 60 to 67%, 55 to 59% and 99 to 105% for epinephrine, normetanephrine and metanephrine, respectively. The prepared cartridge had shown potential and was found robust in extracting the polar analytes repeatedly without any significant loss in efficiency.
Keywords: Silica monolith cartridge; Solid phase extraction; Polar compounds extraction;

In the present work we are presenting a detailed in vitro study of using laser-induced breakdown spectroscopy (LIBS) as a quick and simple method for spectrochemical analysis to identify and characterize some types of human malignancies. This has been performed via detection of the abundance of certain elements namely calcium and magnesium in malignant tissues with respect to the non-neoplastic ones. In order to improve the performance of the LIBS technique – in particular its sensitivity, reproducibility and limit of detection – the measurements have been performed under vacuum (10−2  Torr) and the samples were frozen down to −196 °C in a specially designed vacuum chamber. Under such experimental arrangements a pronounced enhancement has been achieved in the signal to noise (S/N) ratio of different spectral lines. Significant discriminating results have been obtained in case of breast and colorectal cancers indicating the possibility of adopting LIBS in the early detection of the malignancy as well as the identification of the severity and the grade of the disease. The present work demonstrated that future in vivo measurements are also feasible and reliable using especial endoscopic systems for delivery of laser beam and collection of the emitted plasma light.
Keywords: Cancer diagnosis; Laser-induced breakdown spectroscopy; Human malignancies; Breast cancer; Colorectal cancer;

A Cu2O-doped RuO2 sensing electrode (SE) for potentiometric detection of dissolved oxygen (DO) was prepared and its structure and electrochemical properties were analyzed by scanning electron microscopy (SEM), X-ray diffraction (XRD), X-ray photoelectron microscopy (XPS) and energy-dispersive spectroscopy (EDS) techniques. Cu2O-RuO2-SE displayed a linear DO response from 0.5 to 8.0 ppm (log[O2], −4.73 to −3.59) within a temperature range of 9–30 °C. The maximum sensitivity of −47.4 mV/decade at 7.27 pH was achieved at 10 mol% Cu2O. Experimental evaluation of the Cu2O-doped RuO2-SE demonstrated that the doping of RuO2 not only improves its structure but also enhances both sensor's selectivity and antifouling properties. Selectivity measurements revealed that 10 mol% Cu2O-doped RuO2-SE is insensitive to the presence of Na+, Mg2+, K+, Ca2+, NO3 , PO4 2− and SO4 2− ions in the solution in the concentration range of 10−7–10−1  mol/l.
Keywords: Cu2O–RuO2; Water quality sensors; Dissolved oxygen; Thin films;

Hybrid plasma bonding for void-free strong bonded interface of silicon/glass at 200 °C by M.M.R. Howlader; M.G. Kibria; F. Zhang; M.J. Kim (508-515).
A novel hybrid plasma bonding (HPB) that combines sequential plasma activation (reactive ion etching followed by microwave radicals) with anodic bonding has been developed to achieve void-free and strong silicon/glass bonding at low temperature. The interfacial voids were observed at the silicon/glass interface both in the anodic bonding and in the plasma activated anodic bonding, but the voids were completely disappeared in the HPB method at 200 °C. The bonding strength of the silicon/glass in the HPB was as high as 30 MPa at 200 °C, which was higher than that in the individual treatment of anodic and plasma activated bonding methods. The improved characteristic behavior of the interface in the HPB is attributed to the higher hydrophilicity and smooth surfaces of silicon and glass after sequential plasma activation. These highly reactive and clean surfaces enhance the mobility of alkaline cations from the glass surface across the interface toward the bulk of glass in the HPB. This transportation resulted in a ∼353 nm thick alkaline depletion layer in the glass and enlarged the amorphous SiO2 across the interface. The void-free strong bonding is attributed to the clean hydrophilic surfaces and the amorphous SiO2 layer across the interface.
Keywords: Hybrid plasma bonding; Sequential plasma activation; Anodic bonding; Interfacial voids; Bonding strength; Hydrophilicity; Surface roughness; Electrostatic force; High-resolution transmission electron microscopy;

A label-free colorimetric detection of lead ions by controlling the ligand shells of gold nanoparticles by Yu-Lun Hung; Tung-Ming Hsiung; Yi-You Chen; Chih-Ching Huang (516-522).
We have developed a simple, colorimetric and label-free gold nanoparticle (Au NP)-based probe for the detection of Pb2+ ions in aqueous solution, operating on the principle that Pb2+ ions change the ligand shell of thiosulfate (S2O3 2−)-passivated Au NPs. Au NPs reacted with S2O3 2− ions in solution to form Au+·S2O3 2− ligand shells on the Au NP surfaces, thereby inhibiting the access of 4-mercaptobutanol (4-MB). Surface-assisted laser desorption/ionization time-of-flight ionization mass spectrometry (SALDI–TOF MS) and inductively coupled plasma mass spectrometry (ICP-MS) measurements revealed that PbAu alloys formed on the surfaces of the Au NPs in the presence of Pb2+ ions; these alloys weakened the stability of the Au+·S2O3 2− ligand shells, enhancing the access of 4-MB to the Au NP surfaces and, therefore, inducing their aggregation. As a result, the surface plasmon resonance (SPR) absorption of the Au NPs red-shifted and broadened, allowing quantitation of the Pb2+ ions in the aqueous solution. This 4-MB/S2O3 2−–Au NP probe is highly sensitive (linear detection range: 0.5–10 nM) and selective (by at least 100-fold over other metal ions) toward Pb2+ ions. This cost-effective sensing system allows the rapid and simple determination of the concentrations of Pb2+ ions in real samples (in this case, river water, Montana soil and urine samples).
Keywords: Lead ions; Gold nanoparticles; Ligand shell; Label-free; Colorimetric detection;

Study of matrix effects and spectral interferences in the determination of lead in sediments, sludges and soils by SR-ETAAS using slurry sampling by Marianela Savio; Soledad Cerutti; Luis D. Martinez; Patricia Smichowski; Raúl A. Gil (523-527).
An interference-free, fast, and simple method is proposed for Pb determination in environmental solid samples combining slurry sampling and electrothermal atomic absorption spectrometry. Samples were ground to an adequate particle size and slurries were prepared by weighing from 0.05 g to 0.20 g of dry sediment, adding nitric acid, and a solution containing 0.1% Triton X-100. Ultrasonic agitation was employed for slurries homogenization. Analytical variables including acid pre-treatment conditions, particle size, slurry stability, temperature program of the graphite furnace, and type and concentration of the chemical modifier were studied. The undesirable effects of potential non-specific and spectral interferences on Pb signal were also taken into account. Continuum source and self-reversal methods for background correction were evaluated and compared. For calibration, synthetic acid solutions of Pb were employed. Calibration was linear within the range 1–30 μg L−1 and 5–30 μg L−1 when the 217.0 nm and 283.3 nm analytical lines were used. Correlation coefficients of 0.9992 and 0.9997 were obtained. Using optimized conditions, limits of detection (3σ) of 0.025 μg g−1 and 0.1 μg g−1 were achieved for the 217.0 nm and 283.3 nm analytical lines, respectively. The method was successfully applied to the determination of lead in soil, contaminated soil, municipal sludge, and sediment samples. The accuracy was assessed by the analysis of two certified reference materials: municipal sludge (QC MUNICIPAL SLUDGE A) and lake sediment (TRAP-LRM from IJS).
Keywords: Lead; Contaminated soil analysis; Municipal sludge; Lake sediment; Slurry sampling; Self-reversal background corrector; ETAAS;

Magnetic bead based immunoassay for enumeration of CD4+ T lymphocytes on a microfluidic device by Dan Gao; Hai-Fang Li; Guang-Sheng Guo; Jin-Ming Lin (528-533).
Human immunodeficiency virus (HIV) diagnostics are urgently needed in resource-scarce settings. Monitoring of HIV-infected patients requires accurate counting of CD4+ T lymphocytes. However, the current methods for enumeration of CD4+ T lymphocytes are of high cost, technically complex and time-consuming. In this paper, we developed a simple, rapid and inexpensive one-step immunomagnetic method for separating and counting CD4+ T lymphocytes on microfluidic devices with enlarged reaction chambers. CD4+ T lymphocytes were successfully separated and captured from the cell suspension obtained from mouse thymus. CD4 counts were determined under an optical microscope in a rapid and simple format. In order to acquire the maximum efficiency of cell capture, relative parameters were investigated, including section area of the reaction chamber and injection flow rate of the cell suspension. The enlarged reaction chamber with two symmetrical cone-shaped ends was helpful for cell capture, and the maximum capability of captured CD4+ T lymphocytes was about 700 cells μL−1. Our investigations avoided the complex sample pre-treatment, and the entire analysis time was significantly reduced to 15 min. This CD4 counting microdevice had the potential to reduce the cost for HIV diagnosis in resource-limited settings.
Keywords: Microfluidic device; Magnetic beads; Cell separation; CD4+ T lymphocytes enumeration;

This report described the direct voltammetric detection of peroxynitrite (ONOO) at a novel cyanocobalamin modified glassy carbon electrode prepared by electropolymeriation method. The electrochemical behaviors of peroxynitrite at the modified electrode were studied by cyclic voltammetry. The results showed that this new electrochemical sensor exhibited an excellent electrocatalytic activity to oxidation of peroxynitrite. The mechanism of catalysis was discussed. Based on electrocatalytic oxidation of peroxynitrite at the poly(cyanocobalamin) modified electrode, peroxynitrite was sensitively detected by differential pulse voltammetry. Under optimum conditions, the anodic peak current was linear to concentration of peroxynitrite in the range of 2.0 × 10−6 to 3.0 × 10−4  mol L−1 with a detection limit of 1.0 × 10−7  mol L−1 (S/N of 3). The proposed method has been applied to determination of peroxynitrite in human serum with satisfactory results. This poly(cyanocobalamin) modified electrode showed high selectivity and sensitivity to peroxynitrite determination, which could be used in quantitative detection of peroxynitrite in vivo and in vitro.
Keywords: Peroxynitrite; Cyanocobalamin; Chemical modified electrode; Differential pulse voltammetry;

A simple method to synthesize electrostatically self-assembled azides on zinc sulfide nanoparticles (ZnS-N3 NPs) was described and then it was further applied as a multifunctional nanoprobe such as enriching, desalting, accelerating and separation-/washing free nanoprobes for rapid analysis of peptides and proteins and microwave assisted tryptic digested proteins in matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). The ZnS-N3 NPs were characterized by UV–vis, FT-IR, SEM and TEM spectroscopy. The ZnS-N3 NPs can effectively enrich signal intensities for 2–10 times for various peptides and proteins including HW6, insulin, ubiquitin, cytochrome c, lysozyme, myoglobin and bovine serum albumin (BSA) in MALDI-TOF MS. Furthermore, we also demonstrated that the ZnS-N3 NPs can serve as accelerating probes for microwave assisted tryptic digestion of proteins in MALDI-TOF MS. The applicability of the present method on complex sample analysis such as milk proteins from cow milk and ubiquitin and ubiquitin like proteins from oyster mushroom were also demonstrated.
Keywords: ZnS-N3 NPs; Peptides; Proteins; Microwave enzymatic digestion; MALDI-TOF MS;

A ThermoFinnigan sheath liquid flow capillary electrophoresis–mass spectrometry system designed for coupling via a co-axial interface was coupled through an adapted via an alternative, commercially available interface for orthogonal sampling. The affordable, reversible structural alterations made in the commercial LC–MS interface resulted in improved analytical performance.The results of a conventional capillary electrophoresis (CE) method using a commercial co-axial source to determine antioxidant phenolic acids present in virgin olive oil, were compared with those obtained by using a modified orthogonal sampling position. In both cases, separations were done using a 10 mM ammonium acetate/ammonium hydroxide buffer solution at pH 10.0 and a constant applied voltage of 25 kV. The operating variables for the mass spectrometry interface were re-optimized for the modified orthogonal orientation. This allowed the sheath liquid, sheath gas flow rates and capillary voltage to be lowered with respect to the co-axial coupling configuration. In addition, the orthogonal sampling position provided a higher selectivity by effect of ion sampling excluding larger droplets—with an increased momentum along the axis—which were drained through the sink at the bottom of the ion source. Also, the new configuration facilitated sample ionization, improved electrospray stability and led to stronger signals as a result.The new system was validated in terms of precision (repeatability), linearity, and limits of detection and quantification. A comparison of the validation data with the results previously obtained by using a commercial co-axial configuration revealed the adapted orthogonal sampling position to provide better repeatability in both migration times and relative peak areas (<1% and 7% respectively with n  = 15 replicates), a good linear range (with levels in the microgram-per-litre region) and lower limits of detection—especially for the compounds detected with the lowest sensitivity when co-axial ESI was used, as HFA, GEN, FER and VAN finding LOD among 24–3.0 μg L−1 respectively.
Keywords: Antioxidants; CE–ESI-MS; Electrospray source; Orthogonal coupling; Phenolic acids;

Cavity microelectrode for studying manganese dioxide powder as pH sensor by Christine Cachet-Vivier; Bernard Tribollet; Vincent Vivier (555-559).
The potential–pH response of an electrolytic manganese dioxide is investigated by means of a cavity microelectrode (CME). The potential–pH curves show a complex evolution that could be explained by the disporportionation of MnOOH species, leading to the formation of Mn2+ ions on the MnO2 surface. Such a behaviour is not suited for pH sensor application. However when the tip of the electrode is coated by a Nafion membrane, the potential–pH evolution shows a unique slope close to −60 mV pH−1. In addition, the sensor exhibits short time responses to pH variations, a good selectivity, and it can be easily renewed compared to classical sensors.
Keywords: MnO2; pH microsensor; Cavity microelectrode;

A microflow chemiluminescence system for determination of chloramphenicol in honey with preconcentration using a molecularly imprinted polymer by Wisanu Thongchai; Boonsom Liawruangath; Saisunee Liawruangrath; Gillian M. Greenway (560-566).
A novel chemiluminescence (CL) microfluidic system incorporating a molecularly imprinted polymer (MIP) preconcentration step was used for the determination of chloramphenicol in honey samples. The MIP was prepared by using chloramphenicol as the template, diethylaminoethyl methacrylate (DAM) as the function monomer, ethylene glycol dimethacrylate (EGDMA) as the cross-linking monomer, 2, 2′-dimethoxy-2-phenylacetophenone (DMPA) as the free radical initiator and toluene and dodecanol as the solvent. The MIP was pre-loaded into a 10 mm long, 2 mm wide and 150 μm deep channel in a planar glass microfluidic device. When the sample containing chloramphenicol was introduced into the microfluidic device it was first preconcentrated on the MIP then detected by an enhancement effect on the chemiluminescence reaction of tris(2, 2′-bipyridyl) ruthenium(II) with cerium(IV) sulphate in sulphuric acid. A micro-syringe pump was used to pump the reagents. The CL intensity was linear in relationship to the chloramphenicol concentrations from 1.55 × 10−4 to 3.09 × 10−3  μmol L−1 (r 2  = 0.9915) and the detection limit (3σ) and the quantitation limit (10σ) were found to be 7.46 × 10−6 and 2.48 × 10−5  μmol L−1, respectively. This method offered a high selectivity and sensitivity for quantitative analysis of chloramphenicol in the honey samples.
Keywords: Chloramphenicol; Molecularly imprinted polymer (MIP); Chemiluminscence; Microfluidics; MicroFIA;

A method based on Matrix Solid-Phase Dispersion (MSPD) has been developed for the determination of 5 mycotoxins (ochratoxin A and aflatoxins B and G) in different cereals. Several dispersants, eluents and ratios were tested during the optimization of the process in order to obtain the best results. Finally, samples were blended with C18 and the mycotoxins were extracted with acetonitrile. Regarding to matrix effects, the results clearly demonstrated the necessity to use a matrix-matched calibration to validate the method. Analyses were performed by liquid chromatography–triple quadrupole-tandem mass spectrometry (LC–QqQ-MS/MS). The recoveries of the extraction process ranged from 64% to 91% with relative standard deviation lower than 19% in all cases, when samples were fortified at two different concentrations levels. Limits of detection ranged from 0.3 ng g−1 for aflatoxins to 0.8 ng g−1 for OTA and the limits of quantification ranged from 1 ng g−1 for aflatoxins to 2 ng g−1 for OTA, which were below the limits of mycotoxins set by European Union in the matrices evaluated. Application of the method to the analysis of several samples purchased in local supermarkets revealed aflatoxins and OTA levels.
Keywords: Matrix Solid-Phase Dispersion; Aflatoxins; Ochratoxin A; Cereals analysis;

A field flow approach for the in situ monitoring of wastewater quality is developed and assessed in this work, based on a combination of methods employing deconvolution of molecular absorption spectra and in situ/on-line analysis of wastewater effluent of various origin. The approach involves in situ immersion probes to monitor basic physicochemical parameters followed by UV spectrum deconvolution in order to provide a rapid estimate of organic matter, suspended solids and nitrate and on-line analysis of phosphates in a fully automated setup. The collected data are then treated with a series of supervised pattern recognition techniques in order to classify wastewater effluent according to their origin in three major categories namely municipal, industrial and hospital. The results suggest that the method affords a good approximation of realistic concentrations, as determined by reference methods, while it affords a good classification among various wastewater effluents of different origin. In that manner, the method enables a rapid inference of treated wastewater quality and a robust assessment of treatment process state, especially with regards to violations of effluent quality parameters.
Keywords: Classification; In situ monitoring; Pattern recognition techniques; UV spectra deconvolution; Wastewater;

A modified three-step sequential extraction procedure proposed by the Commission of European Communities Bureau of Reference (BCR) was applied to certified reference materials of three different soil groups (rendzina, luvisol, and cambisol) and sewage sludge of different compositions originating from a municipal water treatment plant in order to assess potential mobility and the distribution of molybdenum in the resulting fractions. In the soils examined, molybdenum was present almost entirely in the mineral lattice, the content of molybdenum in the fractions of the studied reference materials of sludges was predominant in the fraction, represents Mo bound to organic matter and sulphide.The internal check of accuracy was performed on the results of the sequential extraction by comparing of the extractable amounts of molybdenum in the sequential procedure with the results of the pseudototal digestion of original samples. The recovery ranged from 96 to 101% and the precision (RSD) in the extracts was below 10%.
Keywords: Molybdenum; ET-AAS; BCR sequential extraction procedure; Soil sludge;

Indirect ultrasonication for protein quantification and peptide mass mapping through mass spectrometry-based techniques by R.J. Carreira; C. Lodeiro; M. Reboiro-Jato; D. Glez-Peña; F. Fdez-Riverola; J.L. Capelo (587-593).
We report in this work a fast protocol for protein quantification and for peptide mass mapping that rely on 18O isotopic labeling through the decoupling procedure. It is demonstrated that the purity and source of trypsin do not compromise the labeling degree and efficiency of the decoupled labeling reaction, and that the pH of the labeling reaction is a critical factor to obtain a significant 18O double labeling. We also show that the same calibration curve can be used for MALDI protein quantification during several days maintaining a reasonable accuracy, thus simplifying the handling of the quantification process. In addition we demonstrate that 18O isotopic labeling through the decoupling procedure can be successfully used to elaborate peptide mass maps. BSA was successfully quantified using the same calibration curve in different days and plasma from a freshwater fish, Cyprinus carpio, was used to elaborate the peptide mass maps.
Keywords: 18O-labeling; Ultrasound; Sonoreactor; Peptide mass mapping; Differential protein expression; MALDI-TOF-MS; Protein quantification;

Approach for rapid extraction and speciation of mercury using a microtip ultrasonic probe followed by LC–ICP-MS by Isabel López; Susana Cuello; Carmen Cámara; Yolanda Madrid (594-599).
A fast method for mercury extraction from biological samples based on the use of HCl leaching plus different enzymatic hydrolysis (with and without mercury complexing agents), and the use of focussed ultrasounds (2-mm microtip) is here proposed. Total mercury content in several biological samples was determined by FI–ICP-MS using a carrier solution consisting of 0.1% (v/v) HCl, 0.1% (v/v) 2-mercaptoethanol, to avoid memory effect, and 0.15% (w/v) KCl. For mercury speciation a RP18 chromatographic column coupled to ICP-MS was used. A mobile phase consisting of 0.1% (v/v) formic acid, 0.1% (v/v) HFBA, 2% (v/v) methanol, and 0.02% (w/v) mM l-cysteine at pH 2.1 was used for chromatographic separation of the mercury species in the sample extracts. Extraction procedures were validated by using 50 mg of tuna fish tissue CRM-463 (2.85 ± 0.16 mg kg−1 for methylmercury). The recoveries obtained were 99 ± 3% and 93 ± 1% after acid leaching (HCl 7 M) and enzymatic extraction (15 mg protease type XIV in 2.5% (v/v) 2-mercaptoethanol), respectively. The optimal sonication conditions (5 min of exposure time and 40% of ultrasound amplitude) were applied to 5 mg of CRM-463 (88 ± 5%), 5 mg of mussel tissue (81 ± 11%) and to 2 mg of zebra fish embryos (90 ± 10%) obtaining good recoveries in all cases. Methylmecury was found to be the most abundant Hg specie in all samples. The developed method is simple and rapid (5 min sample treatment); it is suitable for very small samples and does not alter the original form of the mercury species. Thus, it is of special interest in those cases in which validation of the results may often be hampered by lack of sample availability.
Keywords: Extraction; Ultrasonic probe; 2-Mercaptoethanol; Biological samples; Mercury speciation;

In this paper authors describe a GC-MS acquisition study, relating to the most common, non-steroidal anti-inflammatory drugs (NSAIDs), such as ibuprofen, naproxen, ketoprofen and diclofenac. As novelties to the field, for the trimethylsilyl (TMS) oxime ester derivatives of NSAIDs, at first, a tandem mass spectrometric (MS/MS) acquisition method has been developed, and, also for the first time, the three acquisition techniques, the full scan (FS), the selective ion monitoring (SIM) and the currently optimized MS/MS ones, have been compared: all three in parallel, under strictly the same derivatization/instrumental conditions, both from model solutions and from the Danube River samples. Critical evaluation of the three acquisition protocols was collated on their analytical performances and validated with the same characteristics like the six point calibration curve, the relative standard deviation percentages (RSD%) of parallel tests, the limit of quantitation (LOQ) and the instrumental limit of quantitation (ILQ) values. Data of six point calibration (r2  ≥ 0.997) and RSD% (average: 5.8 RSD%) values proved to be independent on the acquisition methods, while, LOQ and ILQ values furnished considerable differences. Decreasing LOQ data, (expressed in ng/L concentrations) were obtained in the FS, SIM, MS/MS line for ibuprofen (1.0, 0.43, 0.41), naproxen (1.1, 1.0, 0.42), ketoprofen (2.6, 1.0, 0.49) and diclofenac (1.4, 0.41, 0.21), respectively. The same trend was determined in terms of the ILQ values. The practical utility of the optimized MS/MS technique was confirmed by the quantitation of the NSAID contents of the Danube River samples, determined by all three acquisition techniques. Results obtained confirmed the primary importance of the MS/MS acquisition method, even in comparison to the SIM one: avoiding the extreme overestimation of the ibuprofen (≈100%) and ketoprofen (≈400%) concentrations in the Danube River samples.
Keywords: Non-steroidal anti-inflammatory drugs; GC-MS FS; GC-MS SIM; GC-MS/MS; Mass fragmentation; Danube River; Drinking water;

Direct and rapid electrochemical immunosensing system based on a conducting polymer by B. Piro; Q.D. Zhang; S. Reisberg; V. Noel; L.A. Dang; H.T. Duc; M.C. Pham (608-612).
A system device using multifunctional conjugated copolymer poly(5-hydroxy-1,4-naphthoquinone-co-hydroxy-2-thioacetic acid-1,4-naphthoquinone) acting both as immobilizing and transducing element for reagentless immunosensor has been constructed. Its functionality was evaluated in an antigen–antibody interaction model using ovalbumin–anti-ovalbumin. It was shown that the system specifically detects via electrochemical signal the antigen–antibody immune interaction in a reagentless context. Comparison to the conventional ELISA technique relevant to performance and sensitivity was presented.
Keywords: Conjugated polymer; Electrochemical immunosensor; Reagentless detection; Antibody–antigen affinity; ELISA assays;

Discrimination of apple juice and herbal liqueur brands with solid-state electrodes covered with polyaniline and thiacalixarenes by G.A. Evtugyn; S.V. Belyakova; R.V. Shamagsumova; A.A. Saveliev; A.N. Ivanov; E.E. Stoikova; N.N. Dolgova; I.I. Stoikov; I.S. Antipin; H.C. Budnikov (613-619).
Solid-contact ion-selective electrodes based on glassy carbon electrode covered with electropolymerized polyaniline and tetrasubstituted thiacalix[4]arene ionophores with hexyl and o-pyridylamido functional groups at the lower rim have been developed and examined in the discrimination of the brands of apple juices and herbal liqueurs. For this purpose, the liquids tested were diluted and spiked with a constant amount of Fe3+ ions. The variation of the signal toward Fe3+ ions was achieved due to their involvement in the reactions with the organic ligands and the antioxidants present. As was shown, the combination of the three electrodes with various receptors makes it possible to predict the brand of apple juices and herbal liqueurs using linear discriminant analysis in 95–100% cases. The discrimination procedure makes it possible to discriminate liquids within 20 min. Besides, the electrodes developed make it possible to detect individual antioxidants (ascorbic, malic, oxalic acids, hydroquinone, and quercetin) in the range from 5.0 × 10−6 to 1.0 × 10−2  M in direct potentiometric measurements and redox titration.
Keywords: Solid-contact ion-selective electrode; Thiacalixarene; Principal component analysis; Linear discriminant analysis;

Novel luminescent Ir(III) dyes for developing highly sensitive oxygen sensing films by M. Marin-Suarezdel Toro; J.F. Fernandez-Sanchez; E. Baranoff; Md.K. Nazeeruddin; M. Graetzel; A. Fernandez-Gutierrez (620-626).
New sensing films have been developed for the detection of molecular oxygen. These films are based on luminescent Ir(III) dyes incorporated either into polystyrene (with and without plasticizer) or metal oxide, nanostructured material. The preparation and characterization of each film have been investigated in detail. Due to their high sensitivity for low oxygen concentration, the parameters p O 2 ( S = 1 / 2 ) and ΔI 1% have been also evaluated in order to establish the most sensitive membrane for controlling concentrations between 0 and 10% and low oxygen concentrations (lower than 1%), respectively. The results show that the use of nanostructured material increased the sensitivity of the film; the most sensitive membrane for controlling O2 between 0 and 10% is based on N1001 immobilized in AP200/19 (k sv  = 2848 ± 101 bar−1 and p O 2 ( S = 1 / 2 ) = 0.0006 ), and the complex N969 incorporated into AP200/19 seems to be the most suitable for applications in oxygen trace sensing (ΔI 1%  = 93.13 ± 0.13%).
Keywords: Luminescence; Sensor; Iridium complex; Oxygen; Nanostructured material;

Bottom sediments are a very important component of aquatic ecosystems. The sediment matrix is highly diverse and heterogeneous; in consequence, compounds entering the aquatic environment from different sources are considerably enriched at its surface. Bottom sediments are regarded as natural sorbents, since they accumulate many harmful substances, such as heavy metals and stable organic contaminants.Extraction is a key stage in every analytical procedure. It is during this stage that standards are added to samples. Standards are necessary not only for estimating analyte yields but also for validating the whole procedure. The question of the addition of standard substances to sediment samples has not been widely addressed in the subject literature, and yet it is of fundamental importance as regards obtaining reliable results of determinations.This paper describes the results of a study on the effect of standard addition techniques on the results of determination of polychlorinated biphenyls in sediment samples (certified reference material: METRANAL™2—river sediment).
Keywords: Polychlorinated biphenyls; Bottom sediment; Internal standard; Extraction technique;

A high sensitive assay platform based on surface-enhanced Raman scattering for quantification of protease activity by Nazife Nur Yazgan; İsmail Hakkı Boyacı; Erhan Temur; Uğur Tamer; Ali Topcu (631-639).
In this study, a new, sensitive, and rapid assay was developed to quantitatively measure the proteolytic enzyme activity using the surface-enhanced Raman scattering (SERS) probe. Two different shapes of gold nanoparticles, gold nanosphere and nanorod particles were produced. SERS label, comprising self-assembled monolayers (SAMs) of Raman reporter molecule (5,5-Dithiobis (2-Nitrobenzoic acid), DTNB), was coated on the surface of the nanoparticles. Two different SERS-based analysis platforms were designed using gold-coated glass slide and polystyrene microtiter plate. The calibration curves were obtained by plotting the intensity of the SERS signal of symmetric NO2 stretching of DTNB at 1326 cm−1 vs. the protease concentration. The effects of nanoparticle geometry and assay platform on the protease assay were investigated and the best working combination of the parameters was selected as rod shaped SERS probe and gold-coated glass slide. The correlation between the protease activity and SERS signal was found to be linear within the range of 0.1–2 mU/mL (R 2  = 0.979). The limit of detection (LOD) and limit of quantification (LOQ) values of the validated method were found as 0.43 and 1.30 mU/mL, respectively. The intra-day and inter-day precisions of the method, as relative standard deviation (RSD), were determined as 2.5% and 3.6%, respectively. The developed method was successfully applied for quantitative analysis of the commercial enzyme preparate that is used in cheese making process. It was also used for investigation of substrate specificity of protease enzyme towards the casein and bovine serum albumin. The proposed method has a flexibility to try different substrates for the detection of various enzyme activities.
Keywords: Protease activity; SERS; Raman reporter (DTNB); Sphere and rod shaped nanoparticles;

Trans-resveratrol (RVT) is an antioxidant found in red grapes and their derivatives, which has been related to the reduction of cardiovascular diseases and cancer incidence.This work developed a new spectrofluorimetric–chemometric method for the direct determination of RVT in human plasma. For each measurement, excitation–emission matrices were obtained from 280 to 360 nm (excitation) and from 380 to 550 nm (emission). The strategy adopted in this work combined data treatment with parallel factor analysis (PARAFAC), for extracting the pure analyte signal, using the standard addition method, which permits determinations in the presence of a strong matrix effect caused by plasma analyte–protein binding. Plasma samples were diluted 10 times and, for each, four standard additions of RVT were performed, in triplicate. A specific PARAFAC model was built for the three replicates of each sample, from three-way arrays formed by five measurements (initial sample plus four additions), 17 excitation wavelengths and 86 emission wavelengths. The best models were selected with four factors and accounted for more than 99.90% of the data variance. The loadings obtained were related to RVT and three interferences. The scores related to the analyte were used for linear regressions and all standard addition curves presented correlation coefficients equal or greater than 0.99. Good results were obtained in the concentration range from 0.10 to 5.00 μg mL−1, with recoveries between 94.0 and 110.0%. The proposed method was also validated through the estimates of several figures of merit: sensitivity, analytical sensitivity, selectivity, precision, and limits of detection and quantitation.
Keywords: PARAFAC; Trans-resveratrol; Second-order advantage; Chemometrics; Molecular fluorescence; Blood;

A simple and precise square-wave adsorptive cathodic stripping voltammetry (SW-AdCSV) method has been described for simultaneous determination of Mn(II), Cu(II) and Fe(III) in water samples using a carbon paste electrode. In 0.1 mol L−1 acetate buffer (pH 5) containing 50 μmol L−1 of 2-(5′-bromo-2′-pyridylazo)-5-diethylaminophenol (5-Br-PADAP), Mn(II), Cu(II) and Fe(III) were simultaneously determined as metal-complexes with 5-Br-PADAP following preconcentration onto the carbon paste electrode by adsorptive accumulation at +1.0 V (vs. Ag/AgCl/3 M KCl). Insignificant interference from various cations (K+, Na+, Mg2+, Ca2+, Al3+, Bi3+, Sb3+, Se4+, Zn2+, Ni2+, Co2+, Cd2+, Pb2+, V5+, Ti4+ and NH4 +), anions (HCO3 , Cl, NO3−, SO4 2− and PO4 3−) and ascorbic acid was noticed. Limits of detection of 0.066, 0.108 and 0.093 μg L−1 and limits of quantitation of 0.22, 0.36 and 0.31 μg L−1 Mn(II), Cu(II) and Fe(III), respectively, were achieved by the described method. The described stripping voltammetry method was successfully applied for simultaneous determination of Mn(II), Cu(II) and Fe(III) in ground, tap and bottled natural water samples.
Keywords: Manganese; Copper; Iron; 5-Br-PADAP; Determination; Square-wave; Stripping voltammetry;

Using sweeping-micellar electrokinetic chromatography to determine voriconazole in patient plasma by Shu-Chiao Lin; Shu-Wen Lin; Jhong-Min Chen; Ching-Hua Kuo (653-659).
Invasive fungal infection is a life-threatening condition; its occurrence has increased significantly over the past 20 years. We have developed a sensitive and efficient sweeping-micellar electrokinetic chromatography (sweeping-MEKC) method to quantify voriconazole, a potent triazole antifungal drug, in patient plasma. Solid phase extraction (SPE) conditions were first optimized to minimize plasma interference while maintaining a high recovery; the sweeping-MEKC conditions were then systematically optimized to obtain a high sweeping efficiency with good selectivity. Under the optimal analytical conditions, voriconazole was baseline-separated from endogenous materials within 10.5 min with a limit of detection of 0.075 μg mL−1. The background electrolyte comprised 40 mM phosphoric acid, 110 mM sodium dodecyl sulfate, and 20% acetonitrile. In terms of method repeatability, the relative standard deviations (RSDs) of the migration time and the peak area (intra-day; n  = 6) were both less than 5.5%; in terms of intermediate precision, and the RSDs of the peak area and the migration time (inter-day; n  = 3) were both less than 6.3%. We successfully applied this developed method to the quantitative determination of plasma voriconazole levels in 16 patients; the results correlated well with those obtained through analyses using high-performance liquid chromatography. This sweeping-MEKC method is accurate and efficient and appears to be applicable to therapeutic drug monitoring and clinical research.
Keywords: Voriconazole; SPE; Sweeping-MEKC; Plasma;

Bulk optode sensors for batch and flow-through determinations of lead ion in water samples by Chantana Bualom; Wittaya Ngeontae; Sira Nitiyanontakit; Passapol Ngamukot; Apichat Imyim; Thawatchai Tuntulani; Wanlapa Aeungmaitrepirom (660-667).
A sensitive optode consisting of highly lead-selective ionophore (Lead IV), proton-selective chromoionophore (ETH 5294) and lipophilic anionic sites (KTpClPB) in plasticized polyvinyl chloride (PVC) membrane was fabricated. The optode membranes were used for determination of Pb2+ by absorption spectrophotometry in batch and flow-through systems. The influence parameters such as pH, type of buffer solution, response time and concentration of regenerating solution were optimized. The membrane responded to Pb2+ by changing its color from blue to pinkish purple in Tris buffer containing different concentration of Pb2+ at pH 7.0. The optode provided the response range of 3.16 × 10−8 to 5.00 × 10−5  mol L−1 Pb2+ with the detection limit of 2.49 × 10−8  mol L−1 in the batch system within the response time of 30 min. The dynamic range of 1.26 × 10−8 to 3.16 × 10−5  mol L−1 Pb2+ with detection limit of 8.97 × 10−9  mol L−1 were obtained in the flow-through system within the response time of 15 min. Moreover, the proposed optode sensors showed good selectivity towards Pb2+ over Na+, K+, Mg2+, Cd2+, Hg2+ and Ag+. It was successfully applied to determine Pb2+ in real water samples and the results were compared with well-established inductively coupled plasma optical emission spectrometry (ICP-OES). No significant different value (t critical  = 4.30 >  t exp  = 1.00–3.42, n  = 3 at 95% of confidence level) was found.
Keywords: Bulk optode; Lead ion; Lead IV; ETH 5294;

Detection of s-triazine pesticides in natural waters by modified large-volume direct injection HPLC by David J. Beale; Sarit L. Kaserzon; Nichola A. Porter; Felicity A. Roddick; Peter D. Carpenter (668-674).
There is a need for simple and inexpensive methods to quantify potentially harmful persistent pesticides often found in our water-ways and water distribution systems. This paper presents a simple, relatively inexpensive method for the detection of a group of commonly used pesticides (atrazine, simazine and hexazinone) in natural waters using large-volume direct injection high performance liquid chromatography (HPLC) utilizing a monolithic column and a single wavelength ultraviolet–visible light (UV–vis) detector. The best results for this system were obtained with a mobile phase made up of acetonitrile and water in a 30:70 ratio, a flow rate of 2.0 mL min−1, and a detector wavelength of 230 nm. Using this method, we achieved retention times of less than three minutes, and detection limits of 5.7 μg L−1 for atrazine, 4.7 μg L−1 for simazine and 4.0 μg L−1 for hexazinone. The performance of this method was validated with an inter-laboratory trial against a National Association of Testing Authorities (NATA) accredited liquid chromatography–mass spectrometry/mass spectrometry (LC–MS/MS) method commonly used in commercial laboratories.
Keywords: Atrazine; Direct injection HPLC; Large-volume injection; Monolithic column; Hexazinone; Natural organic matter; Natural waters; Simazine;

Individual and simultaneous determination of lead, cadmium, and zinc by anodic stripping voltammetry at a bismuth bulk electrode by Kristie C. Armstrong; Clarissa E. Tatum; Royce N. Dansby-Sparks; James Q. Chambers; Zi-Ling Xue (675-680).
A bismuth bulk electrode (BiBE) has been investigated as an alternative electrode for the anodic stripping voltammetric (ASV) analysis of Pb(II), Cd(II), and Zn(II). The BiBE, which is fabricated in-house, shows results comparable to those of similar analyses at other Bi-based electrodes. Metal accumulation is achieved by holding the electrode potential at −1.4 V (vs. Ag/AgCl) for 180 s followed by a square wave voltammetric stripping scan from −1.4 to −0.35 V. Calibration plots are obtained for all three metals, individually and simultaneously, in the10–100 μg L−1 range, with a detection limit of 93, 54, and 396 ng L−1 for Pb(II), Cd(II), Zn(II), respectively. A slight reduction in slope is observed for Cd(II) and Pb(II) when the three metals are calibrated simultaneously vs. individually. Comparing the sensitivities of the metals when calibrated individually vs. in a mixture reveals that Zn(II) is not affected by stripping in a mixture. However, Pb(II) and Cd(II) have decreasing sensitivities in a mixture. The optimized method has been successfully used to test contaminated river water by standard addition. The results demonstrate the ability of the BiBE as an alternative electrode material in heavy metal analysis.
Keywords: Anodic stripping voltammetry; Bismuth bulk electrode; Lead; Cadmium; Zinc; Heavy metal;

Assemblies of brilliant cresyl violet to DNA in the presence of γ-cyclodextrin by Jin-Ping Song; Yu-Jing Guo; Qiang Zhao; Shao-Min Shuang; Chuan Dong; Martin M.F. Choi (681-686).
The interactions of brilliant cresyl violet (BCV) with herring sperm DNA in γ-cyclodextrin (γ-CD) supramolecular system were studied by UV-vis absorption spectroscopy and cyclic voltammetry (CV). Both UV-vis absorption and CV data show that the interaction of BCV with DNA depends on the concentration ratio of BCV to DNA (R), the initial concentration of BCV and γ-CD. The binding constants of BCV monomer, (BCV)2 dimer and (BCV)2-γ-CD inclusion complex with DNA are 1.64 × 105, 2.56 × 104 and 2.32 × 103  M−1, respectively. It was observed that γ-CD can affect the interactive mode of BCV with DNA. If R is larger than 0.5, the (BCV)2-γ-CD inclusion complex will retain intact and bind to DNA via the electrostatic attraction forces. By contrast, when R is smaller than 0.5, the inclusion complex will be partially dissociated and the free BCV monomer is intercalated into the double-helix structure of DNA attributing to the more favorable microenvironment of DNA for the BCV monomer. Our work postulates the importance of the initial concentration of dye and host molecule on the interaction of dye with DNA in living bodies.
Keywords: Brilliant cresyl violet; γ-Cyclodextrin; DNA; Inclusion complex;

Direct colorimetric visualization of mercury (Hg2+) based on the formation of gold nanoparticles by Aiping Fan; Ying Ling; Choiwan Lau; Jianzhong Lu (687-692).
It is critical to be able to detect and quantify Hg2+ ions under aqueous conditions with high sensitivity and selectivity. The technique presented herein provides a direct way for simple colorimetric visualization of Hg2+ ions in aqueous solution, based on the formation of gold nanoparticles through the Hg2+ catalyzed HAuCl4/NH2OH reaction. The outstanding selectivity and sensitivity result from the well-known amalgamation process that occurs between mercury and gold. The entire procedure takes less than 20 min. The limit of detection (2 ppb) shows excellent potential for monitoring ultralow levels of mercury in water samples.
Keywords: Gold nanoparticle; Mercury; colorimetric detection; HAuCl4; NH2OH;

Direct Determination of Urinary Lysozyme Using Surface Plasmon Resonance Light-Scattering of Gold Nanoparticles by Xinyi Wang; Yao Xu; Xiao Xu; Ke Hu; Minghui Xiang; Limei Li; Feng Liu; Na Li (693-697).
The purpose of this study was to establish a simple and sensitive analytical method for lysozyme using Plasmon Resonance Light-Scattering (PRLS) technique with Gold Nanoparticles (AuNPs) as the probe. Nanomolar level of lysozyme induced AuNPs aggregation with enhanced PRLS. For 1.4 nM citrate-capped AuNPs (13 nm in diameter), the linear range of the calibration curve was 15-50 nM with a detection limit of 13.1 nM for lysozyme. Six nanomolar lysozyme can produce an observable PRLS enhancement. Most potential interfering substances present in urine had a negligible effect on the determination. The interference from human serum albumin in the urinary sample can be reduced by precipitating the albumin with ethanol at pH 4.8-4.9. The 90.1-118.2% recovery was achieved for 8 individual lysozyme-spiked urinary samples. This simple and sensitive method for lysozyme does not require sample clean-up and AuNPs modification, thus provided an alternative for urinary lysozyme determination.
Keywords: Lysozyme; Plasmon Resonance Light-Scattering; Gold Nanoparticles; Urinary sample;

A new method was developed for simultaneous determination of cypermethrin and permethrin residues in pear juice with ultrasound-assisted dispersive liquid-liquid microextraction (UA-DLLME) and gas chromatography-flame ionization detection (GC-FID). 3.5 mL of methanol (dispersant) and 30 μL of C2Cl4 (extractant) were injected into 5.0 mL of pear juice sample and then emulsified by ultrasound for 2.0 min to forming the cloudy solution. Under the optimum condition, the enrichment factors for cypermethrin and permethrin were 344 and 351 fold respectively. Good linearity was observed in a range of 0.009-1.52 μg g−1 with the correlation coefficient (r 2)≥0.9993. The limits of detection (LODs) were 3.1 and 2.2 μg kg−1 for cypermethrin and permethrin, respectively (S/N = 3). The recoveries of the method evaluated at three spiked levels were in the range of 92.1%-107.1%. The repeatability evaluated as intra-day and inter-day precision (RSDs) were less than 4.0% (n = 5). The developed method was successfully applied to determine the two pesticide residues in different pear juice samples.
Keywords: Ultrasound-assisted dispersive liquid-liquid microextraction; Cypermethrin; Permethrin; Gas chromatography; Pear juice samples;

A novel antibody immobilization and its application in immunoaffinity chromatography by Shuoyao Zhang; Junqing Wang; Dawei Li; Juan Huang; Hong Yang; Anping Deng (704-709).
A novel antibody immobilization and its application in immunoaffinity chromatography (IAC) were presented. Using acrylamide (AM) as monomer, ethylene glycoldimethacrylate (EGDMA) as cross-linker and bulk polymerization as synthetic method, we prepared a polymer in which the Cu(II) was embedded. The Cu(II)-embedded polymer was tested for its binding with protein. It was found that Cu(II)-embedded polymer displayed a strong binding with bovine serum albumin (BSA). At 80% of methanol, no BSA was released from Cu(II)-embedded polymer. The Cu(II)-embedded polymer was then used as a novel solid support for antibody immobilization. IAC column was prepared by immobilizing polyclonal antibody (pAb) against clenbuterol (CL) on Cu(II)-embedded polymer and packing the Cu(II)-embedded polymer-pAb into a common solid phase extraction (SPE) cartridge. Under optimal extraction conditions, the IAC column was characterized in terms of maximum binding capacity for target analyte, extraction efficiency and reusability. It was revealed that, for IAC column packed with 0.1 g of solid support immobilized with antibody, the maximum capacity for CL was 616 ng; the extraction recoveries of the column for CL from three spiked food samples were 84.4-95.2% with relative standard deviation (RSD) of 9.3-15.5%; after more than 30 times repeated usage, there was not significant loss of specific recognition. The results demonstrated the feasibility of the prepared IAC column for CL extraction. The proposed antibody immobilization method exhibiting the properties of simplicity, low cost, strong binding for target analyte, no leaching of antibody, etc., would be a very useful tool applied in the field of IAC.
Keywords: antibody immobilization; immunoaffinity chromatography; Cu(II)-embedded polymer; solid phase extraction; clenbuterol;

Determination of endothelin-1 in rats using a high-performance liquid chromatography coupled to electrospray tandem mass spectrometry by Maria Walczak; Andrzej Fedorowicz; Stefan Chłopicki; Joanna Szymura-Oleksiak (710-718).
A sensitive and specific liquid chromatography tandem mass spectrometry method with electrospray ionization for the determination of endothelin-1 in rat plasma and lung effluents has been developed and validated. Detection was achieved by an Applied Biosystems MDS Sciex API 2000 triple quadrupole mass spectrometer coupled to an Agilent 1100 LC system. The limit of detection and the limit of the quantification of ET-1 in matrix buffer was estimated at 40 pM and 1 nM, respectively. The precision and accuracy for both intra- and inter-day determination of the analyte ranged from 2.5% to 14.7% and from 104.2% to 113.3%, respectively. No significant relative matrix effect was observed. Stability of ET-1 established in a bench-top, autosampler, long-term storage stability as well as freeze/thaw cycles shown no significant degradation products in the samples. The results of the method validation indicated that this method is applicable for the determination of the ET-1 concentration in an effluent from the isolated lung preparation as well as in vivo in plasma samples to evaluate ET-1 as a potential biomarker of the progression of pulmonary endothelial dysfunction and pulmonary hypertension in rats induced by a monocrotaline injection.
Keywords: Endothelin-1; Mass spectrometry; Method validation; Pulmonary hypertension; Rats;

Comparative study of solvent extraction and thermal desorption methods for determining a wide range of volatile organic compounds in ambient air by Noelia Ramírez; Anna Cuadras; Enric Rovira; Francesc Borrull; Rosa Maria Marcé (719-727).
This paper compares two analytical methods for determining levels of 90 volatile organic compounds (VOCs) commonly found in industrial and urban atmospheres. Both methods are based on two official methods for determining benzene levels and involve collecting samples by active adsorptive enrichment on solid sorbents. The first method involves solvent extraction and uses activated charcoal as a sorbent. After sampling, the sorbent is extracted with 1 mL of carbon disulfide and then 1 μL of the extract is analysed in a GC–MS. The second method involves thermal desorption (TD) and uses Tenax TA and Carbograph 1TD as sorbents, which allows the whole sample to be analysed. In general, the thermal desorption method showed the best repetitivity and recovery and the lowest limit of detection and quantification for all target compounds. Because of its lower sensitivity, the solvent extraction method needs the preconcentration of large sample volumes of air (720 L vs. 2.64 L for the thermal desorption method) to yield similar limits of detection.The performance of both methods in real samples was tested in a location near to a petrochemical complex. The results of the 24-h samples for the solvent extraction method were compared with the average of 12 2-h samples for the TD method. In some cases, both methods found differences in the VOC concentrations, especially in those compounds whose concentrations fluctuate significantly during the day.
Keywords: Thermal desorption; Solvent desorption; Comparison of methods; Air analysis; Volatile organic compounds;

A highly sensitive and selective dimethyl ether sensor based on cataluminescence by Runkun Zhang; Xiaoan Cao; Yonghui Liu; Yan Peng (728-732).
A sensor for detecting dimethyl ether was designed based on the cataluminescence phenomenon when dimethyl ether vapors were passing through the surface of the ceramic heater. The proposed sensor showed high sensitivity and selectivity to dimethyl ether at an optimal temperature of 279 °C. Quantitative analysis were performed at a wavelength of 425 nm, the flow rate of carrier air is around 300 mL/min. The linear range of the cataluminescence intensity versus concentration of dimethyl ether is 100–6.0 × 103  ppm with a detection limit of 80 ppm. The sensor response time is 2.5 s. Under the optimized conditions, none or only very low levels of interference were observed while the foreign substances such as benzene, formaldehyde, ammonia, methanol, ethanol, acetaldehyde, acetic acid, acrolein, isopropyl ether, ethyl acetate, glycol ether and 2-methoxyethanol were passing through the sensor. Since the sensor does not need to prepare and fix up the granular catalyst, the simple technology reduces cost, improves stability and extends life span. The method can be applied to facilitate detection of dimethyl ether in the air. The possible mechanism of cataluminescence from the oxidation of dimethyl ether on the surface of ceramic heater was discussed based on the reaction products.
Keywords: Cataluminescence; Dimethyl ether; DME; Gas sensor;

A cataluminescence gas sensor based on nanosized V2O5 for tert-butyl mercaptan by Huili Zhang; Lichun Zhang; Jing Hu; Pingyang Cai; Yi Lv (733-738).
This work proposed a gas sensor for the determination of tert-butyl mercaptan, one of the highly toxic volatile sulfur compounds, which was based on cataluminescence emission during its catalytic oxidation on the surface of nanosized V2O5. The cataluminescence characteristics and the optimum conditions, including the morphology of sensing material, the wavelength of cataluminescence emission, the oxygen flow rate and working temperature were investigated in detail. Under the optimized conditions, the calibration curve of the relative cataluminescence intensity versus the concentration of tert-butyl mercaptan vapor was made, with the linear range of 5.6–196 μg mL−1 and the detection limit of 0.5 μg mL−1 (S/N  = 3). The relative standard deviation (R.S.D.) (n  = 5) of relative cataluminescence intensity for 84 μg mL−1 tert-butyl mercaptan was 3.6%. There is no or weak response to some common substances, such as formic acid, alcohol (methanol, ethanol, propanol, isopropanol, n-butanol, isoamyl alcohol), o-dichlorobenzene, acetonitrile, ethyl acetate, aldehyde (formaldehyde, acetaldehyde and propanal), 1,2-dichloroethane and ammonia. Furthermore, the proposed sensor was successfully used for determining tert-butyl mercaptan in four artificial samples, with a good recovery. The results demonstrated that the proposed gas sensor had a promising capability for the tert-butyl mercaptan in routine monitoring.
Keywords: Cataluminescence; Gas sensor; Vanadium pentoxide; Tert-butyl mercaptan;

A new dispersive vapor extraction (DVE) technique for rapid removal of selected volatile organic compounds (VOCs) from gaseous mixtures was investigated. In this technique, less than 1.0 mL of a volatile solvent was vaporized for 8 min in a 250-mL flask containing a gaseous mixture. The flask was then cooled under running tap water for 2–3 min to induce condensation of the vapor and co-extraction of the VOCs from the headspace. The technique was tested over a concentration range of 4–23 ppb, and resulted in extraction efficiencies ranging from 80 to 97% for the VOCs tested. Because of its simplicity and the relatively short sampling time, DVE could potentially lead to high sample throughput and rapid air analysis.
Keywords: VOCs; Extraction; Pre-concentration;

Towards the identification of plant and animal binders on Australian stone knives by Alisa J. Blee; Keryn Walshe; Allan Pring; Jamie S. Quinton; Claire E. Lenehan (745-750).
There is limited information regarding the nature of plant and animal residues used as adhesives, fixatives and pigments found on Australian Aboriginal artefacts. This paper reports the use of FTIR in combination with the chemometric tools principal component analysis (PCA) and hierarchical clustering (HC) for the analysis and identification of Australian plant and animal fixatives on Australian stone artefacts. Ten different plant and animal residues were able to be discriminated from each other at a species level by combining FTIR spectroscopy with the chemometric data analysis methods, principal component analysis (PCA) and hierarchical clustering (HC). Application of this method to residues from three broken stone knives from the collections of the South Australian Museum indicated that two of the handles of knives were likely to have contained beeswax as the fixative whilst Spinifex resin was the probable binder on the third.
Keywords: FTIR; Principle component analysis; Resins; Stone artefacts;

A sensitive, rapid ferricyanide-mediated toxicity bioassay developed using Escherichia coli by Kylie Catterall; David Robertson; Sian Hudson; Peter R. Teasdale; David T. Welsh; Richard John (751-757).
A need for rapid toxicity techniques has seen recent research into developing new microbiological assays and characterising their toxicity responses using a range of substances. A microbiological bioassay that determines changes in ferricyanide-mediated respiration for toxicity measurement (FM-TOX) shows particular promise. The development and optimisation of an improved FM-TOX method, incorporating novel features, is described using Escherichia coli as the biocatalyst. Omission of an exogenous carbon source, used in previously described FM-TOX assays, substantially improves the assay sensitivity. In addition, the development of a two-step procedure (toxicant exposure followed by determination of microbial respiratory activity) was found to enhance the inhibition of E. coli by 3,5-dichlorophenol and four other toxicants, compared to single-step procedures. Other assay parameters, such as the ferricyanide concentration, exposure times and optical density of the biocatalyst were also optimised, sometimes based on practical aspects. Toxicity tests were carried out using the adopted technique on both organic and inorganic toxicants, with classic sigmoid-shaped dose–response curves observed, as well as some non-standard responses. IC50 data is presented for a number of common toxicants. The optimised assay provides a good foundation for further toxicity testing using E. coli, as well as the potential for expanding the technique to utilise other bacteria with complementary toxicity responses, thereby allowing use of the assay in a range of applications.
Keywords: Microbial; Ferricyanide; IC50; Chronoamperometry; Respiration; Inhibition;

Headspace single-drop microextraction has been used for the determination of cyanide with ninhydrin in combination with fibre-optic-based cuvetteless microspectrophotometry which accommodates sample volume of 1 μL placed between the two ends of optical fibres, and has been found to avoid salient drawbacks of batch methods. This method involved hydrocyanic acid formation in a closed vial, and simultaneous extraction and reaction with 2 μL drop of ninhydrin in carbonate medium suspended at the tip of a microsyringe needle held in the headspace of the acidified sample solution. The method was linear in range 0.025–0.5 mg L−1 of cyanide. The headspace reaction was free from the interference of substances, e.g., thiocyanate, hydrazine sulphate, hydroxylammonium chloride and ascorbic acid. Sulphide was masked by cadmium sulphate, nitrite by sulphamic acid, sulphite by N-ethylmaleimide, and halogens by ascorbic acid. The limit of detection was found to be 4.3 μg L−1 of cyanide which was comparable to existing most sensitive methods for cyanide. However, the present method is far more simple. The method was applied to acid-labile and metal cyanides complexes by treatment with sulphide when metal sulphides were precipitated setting cyanide ion free, and to iron(II) and (III) cyanide complexes by their decomposition with mercury(II), the mercury(II) cyanide formed was then determined. These pre-treatment methods avoided cumbersome pre-separation of cyanide by methods such as distillation or gas diffusion. The overall recovery of cyanide in diverse samples was 97% with RSD of 3.9%.
Keywords: Microspectrophotometry; Fibre optics; Cuvetteless; Headspace single-drop microextraction; Free and total cyanide; Ninhydrin;

A novel method for electrically assisted microextraction coupled to liquid chromatography–mass spectrometry was evaluated for determination of trace levels of parathion in water. A pencil lead electrode was used in a di-electrode system to extract parathion onto the electrode surface with a reductive potential applied. The optimum extraction conditions were found to be a potential of −600 mV for 60 s in pH 2 phosphate buffer solution. The parathion was desorbed statically for 1 min and dynamically for 3 min in the commercial SPME–HPLC desorption chamber, then analyzed with LC–APCI–MS/MS. The detection limit (LOD) for parathion in water was found to be 0.3 ng/mL. The proposed technique was demonstrated to be fast, sensitive and not require a solvent sample pretreatment.
Keywords: Electrically assisted solid-phase microextraction; Parathion; Liquid chromatography–mass spectrometry;

Fluorescence resonance energy transfer (FRET) is widely used to obtain the distance between a donor and an acceptor in biological research. However, the detection of FRET efficiencies with fluorescence microscopy imaging systems remains a great challenge due to the difficulties of transferring gray scales of the images into fluorescence intensities, and the absence of exact quantum yields of donors and acceptors. Herein, we presented a new method to detect the FRET efficiency in imaging systems by analyzing the photo-bleaching-induced changes in fluorescent intensities of quantum dots (QDs, donors) and Cy5 dyes (acceptors). Our method is different from the previous acceptor-photo-bleaching studies in imaging systems by theoretically analyzing the bleaching process, and bringing forward a new parameter which is universal for samples of the same kind. It is convenient for calculating FRET efficiencies. There is hardly any spectral crosstalk between 605QD and Cy5, thus the FRET result is more accurate than that of many other common FRET pairs. The lengths of single-stranded and double-stranded DNA fragments in solution were determined via the analysis of FRET efficiency values. This technique provides a reliable approach to study biomacromolecules in living cells through fluorescent imaging and in situ measurements.
Keywords: FRET efficiency; Microscopy imaging; Photo-bleaching; Quantum dot;

Different substituent groups were introduced onto the rim of β-cyclodextrin through rigid C=N bonds to form a series of imino-modified β-cyclodextrin derivatives: mono(6-deoxy-phenylimino)-β-cyclodextrin (BCD), mono(6-deoxy-isopropylimino)-β-cyclodextrin (YBCD), mono(6-deoxy-N-1-phenylethylimino)-β-cyclodextrin (R-,S-BYCD), mono[6-deoxy-N-1-(2-hydroxyl)-phenylethylimino]-β-cyclodextrin (R-,S-PGCD), heptakis(2,6-o-diamyl-6-deoxy-phenylimino)-β-cyclodextrin (WBCD), heptakis(2,6-o-diamyl-6-deoxyisopropylimino)-β-cyclodextrin (WYBCD) and heptakis[2,6-o-diamyl-6-deoxy-R-(-)-N-1-phenylethylimino)-β-cyclodextrin (WRBYCD). The obtained derivatives were then bonded to silica gel and used in high-performance liquid chromatography (HPLC) as chiral stationary phases (CSPs). The separation performance of these CSPs was examined by separating disubstituted benzenes, amino acids, ferrocene derivatives andchiral aromatic alcohol compounds. Satisfactory separation results were obtained for most of the compounds. The values for selectivity factors can reach up to 8.50 and 8.16 for separating positional isomers and ferrocene derivatives, respectively, and the best resolution was 6.89 for aromatic alcohol derivative separations. Molecular dynamics (MD) simulations were carried out for chiral discrimination of rac-N-benzoyl-phenylglycinol on S-PGCD CSP to study the recognition mechanism. MD simulation results show that the average free-energy of interaction is −1304.83 kcal/mol for the l-enantiomer and S-PGCD and −1324.23 kcal/mol for the d-enantiomer and S-PGCD. In the recognition stage, the l-enantiomer moves along the exterior of the cyclodextrin cavity from the wider edge to the narrower edge of cyclodextrin whereas the d-enantiomer moves slightly towards the cavity. The l-enantiomer thus is separated first due to weaker interaction with S-PGCD.
Keywords: Chiral stationary phase; β-cyclodextrin derivatives; HPLC; Separation performance; Recognition mechanism;

A novel and sensitive extraction procedure using maghemite nanoparticles (γ-Fe2O3) modified with sodium dodecyl sulfate (SDS), as an efficient solid phase, was developed for removal, preconcentration and spectrophotometric determination of trace amounts of malachite green (MG) and leuco-malachite green (LMG). Combination of nanoparticle adsorption and easily magnetic separation was used to extraction and desorption of MG and LMG. The adsorption capacity was evaluated using both the Langmuir and Freundlich adsorption isotherm models. Maghemite nanoparticles were prepared by co-precipitation method and their surfaces were modified by SDS. The size and properties of the produced maghemite nanoparticles was determined by X-ray diffraction (XRD) analysis, scanning electron microscopy (SEM) and BET analysis. MG and LMG became adsorbed at pH 3.0. LMG was oxidized to MG by adsorption on maghemite nanoparticles. The adsorbed MG was then desorbed and determined spectrophotometrically. The calibration graph was linear in the range 0.50–250.00 ng mL−1 of MG and LMG with a correlation coefficient of 0.9991. The detection limit of the method for determination of MG was 0.28 ng mL−1 and the relative standard deviation (R.S.D.) for 10.00 and 50.00 ng mL−1 of malachite green was 1.60% (n  = 3) and 0.86% (n  = 5), respectively. A preconcentration factor of 50 was achieved in this method. The Langmuir adsorption capacity (q max) was found to be 227.3 mg g−1 of the adsorbent. The method was applied to the determination of MG in fish farming water samples.
Keywords: Malachite green; Leuco-malachite green; Spectrophotometric determination; Maghemite nanoparticles; Preconcentration;

A sensitive and robust analytical method for spectrophotometric determination of ethyl xanthate, CH3CH2OCS2 at trace concentrations in pulp solutions from froth flotation process is proposed. The analytical method is based on the decomposition of ethyl xanthate, EtX, with 2.0 mol L−1 HCl generating ethanol and carbon disulfide, CS2. A gas diffusion cell assures that only the volatile compounds diffuse through a PTFE membrane towards an acceptor stream of deionized water, thus avoiding the interferences of non-volatile compounds and suspended particles. The CS2 is selectively detected by UV absorbance at 206 nm (ɛ  = 65,000 L mol−1  cm−1). The measured absorbance is directly proportional to EtX concentration present in the sample solutions. The Beer's law is obeyed in a 1 × 10−6 to 2 × 10−4  mol L−1 concentration range of ethyl xanthate in the pulp with an excellent correlation coefficient (r  = 0.999) and a detection limit of 3.1 × 10−7  mol L−1, corresponding to 38 μg L−1. At flow rates of 200 μL min−1 of the donor stream and 100 μL min−1 of the acceptor channel a sampling rate of 15 injections per hour could be achieved with RSD < 2.3% (n  = 10, 300 μL injections of 1 × 10−5  mol L−1  EtX). Two practical applications demonstrate the versatility of the FIA method: (i) evaluation the free EtX concentration during a laboratory study of the EtX adsorption capacity on pulverized sulfide ore (pyrite) and (ii) monitoring of EtX at different stages (from starting load to washing effluents) of a flotation pilot plant processing a Cu–Zn sulfide ore.
Keywords: Ethyl xanthate; Flow analysis; Gas-diffusion; Spectrophotometry; Flotation process; Mining industry;

Voltammetric behavior of naratriptan and its determination in tablets by C. Velasco-Aguirre; A. Álvarez-Lueje (796-802).
The electrochemical behavior and the analytical application of the selective serotonin agonist naratriptan (N-methyl-3-(1-methyl-4-piperidyl)indole-5-ethanesulfonamide) are presented herein. Naratriptan exhibits an anodic response in aqueous media over a broad pH range (pH 2–12), as determined by differential pulse voltammetry and cyclic voltammetry using glassy carbon electrodes. This response is irreversible in nature, diffusion-controlled and probably caused by the oxidation of the naratriptan indole moiety. The differential pulse voltammetry technique was performed in 0.1 mol L−1 Britton–Robinson buffer (pH = 3), which elicited the most reproducible results. The percentage of naratriptan recovery was 102.1 ± 1.8%, and the limits of detection and quantitation were 9.5 × 10−6 and 2.0 × 10−5  mol L−1, respectively. Selectivity trials revealed that the oxidation signal of the drug was not disturbed by the presence of excipients or degradation products. Thus, we conclude that the method presented herein is useful for the quantification of naratriptan in pharmaceutical drugs and that this method requires no separations or extractions. Finally, this voltammetric method was successfully applied to determine the quantity and the content uniformity of naratriptan in drug tablets. A comparison of this technique to the standard high-performance liquid chromatography technique was conducted at the end of our study.
Keywords: Naratriptan; Differential pulse voltammetry; Pharmaceutical;

Label-free optical diagnosis of hepatitis B virus with genetically engineered fusion proteins by Shun Zheng; Do-Kyun Kim; Tae Jung Park; Seok Jae Lee; Sang Yup Lee (803-809).
A simple biosensing strategy for the diagnosis of patients with hepatitis B virus (HBV) was developed. This study can be divided into two themes, both of which utilized gold-binding polypeptide (GBP) fusion proteins: HBV surface antigen PreS2 (HBsAg) detection with GBP-fused single chain antibody (GBP-ScFv) and anti-HBsAg detection with GBP-HBsAg. These GBP-fusion proteins can directly bind onto the gold surface via the high binding affinity between the GBP and the gold surface, while at the same time, orient the recognition sites toward the sample for target binding. This one-step immobilization strategy, which greatly simplifies a fabrication process as well as maintaining biological activity of the recognition elements, can be applied to optical analytical methods, such as surface plasmon resonance (SPR) and localized surface plasmon resonance (LSPR).
Keywords: Surface plasmon resonance; Localized surface plasmon resonance; Biosensor; Hepatitis B virus; Gold-binding polypeptide; Fusion protein;

Surface plasmon resonance immunosensor for bacteria detection by H. Baccar; M.B. Mejri; I. Hafaiedh; T. Ktari; M. Aouni; A. Abdelghani (810-814).
This work describes an approach for the development of two bacteria biosensors based on surface plasmon resonance (SPR) technique. The first biosensor was based on functionalized gold substrate and the second one on immobilized gold nanoparticles. For the first biosensor, the gold substrate was functionalized with acid–thiol using the self-assembled monolayer technique, while the second one was functionalized with gold nanoparticles immobilized on modified gold substrate. A polyclonal anti-Escherichia coli antibody was immobilized for specific (E. coli) and non-specific (Lactobacillus) bacteria detection. Detection limit with a good reproducibility of 104 and 103  cfu mL−1 of E. coli bacteria has been obtained for the first biosensor and for the second one respectively. A refractive index variation below 5 × 10−3 due to bacteria adsorption is able to be detected. The refractive index of the multilayer structure and of the E. coli bacteria layer was estimated with a modeling software.
Keywords: SPR; Biofunctionalization; Immunosensor; Gold nanoparticles;

Derivatization of GSSG by pHMB in alkaline media. Determination of oxidized glutathione in blood by Valeria Angeli; Huilun Chen; Zoltan Mester; Yulan Rao; Alessandro D’Ulivo; Emilia Bramanti (815-820).
Chromatographic determination of glutathione disulfide (GSSG) without any preliminary reduction has been presented using GSSG derivatization by p-hydroxymercuribenzoate (pHMB) in strong alkaline medium followed by the determination of GS–pHMB complex by reversed phase chromatography coupled to chemical vapour generation and atomic fluorescence detector (RPC–CVGAFS). A detection limit of 35 nM for GSSG (corresponding to 1.8 pmol) detected as GS–pHMB species was achieved based on a signal-to-noise ratio of 3 in buffer and in blood. The proposed method was applied to the determination of GSSG in whole blood and validated by the classical determination of GSSG by derivatization after reduction with dithiothreitol (DTT).
Keywords: Oxidized glutathione; GSSG; Liquid chromatography; Atomic fluorescence spectrometry;

Analytical approaches for Hg determination in wastewater samples by means of total reflection X-ray fluorescence spectrometry by E. Marguí; P. Kregsamer; M. Hidalgo; J. Tapias; I. Queralt; C. Streli (821-827).
At present, there is a considerable interest in Hg monitoring in wastewater samples due to its widespread occurrence and the high toxicity of most of its compounds. Hg determination in water samples by means of total reflection X-ray fluorescence spectrometry (TXRF) entails some difficulties due to the high vapor pressure and low boiling point of this element that produce evaporation and loss of Hg from the surface of the reflector during the drying process, commonly used for sample preparation in TXRF analysis.The main goal of the present research was to develop a fast and simple chemical strategy to avoid Hg volatilization during the analysis of wastewater samples by TXRF spectrometry. Three different analytical procedures were tested for this purpose: (i) increasing the viscosity of the wastewater sample by adding a non-ionic surfactant (Triton® X-114), (ii) Hg immobilization on the quartz reflectors using the extractant tri-isobutylphosphine (Cyanex 471X) and (iii) formation of a stable and non-volatile Hg complex into the wastewater sample. The best analytical strategy was found to be the formation of a Hg complex with thiourea (pH = 10) before the deposition of 10 μL of sample on the reflector for following TXRF analysis. Analytical figures of merit such as linearity, limits of detection, accuracy and precision were carefully evaluated. Finally, the developed methodology was applied for the determination of Hg in different types of wastewater samples (industrial effluents, municipal effluents from conventional systems and municipal effluents from constructed wetlands).
Keywords: Hg; TXRF; Wastewater samples;

A simple, precise and accurate automatic method for the determination of total iodine in milk products by indirect atomic absorption spectrometry is proposed. Iodide in solutions resulting from alkaline ashing of samples is precipitated with silver ion in a precipitation–dissolution flow manifold, which allows performing on-line the retention of the silver iodide precipitate formed on a filter, its wash with diluted ammonia and its dissolution with a diluted thiosulfate solution. Dissolved silver is also determined on-line by flame atomic absorption, and the achieved amount of this metal is proportional to that of iodine in the sample. The proposed method is very selective, avoids interferences from anions present in the samples, which can be also precipitated with silver, because these silver compounds are dissolved with ammonia at the washing step. This method allows the determination of iodine in the range 0.011–0.35 μg mL−1 with a relative standard deviation between 1.3 and 6.8% at a rate of ca. 17 samples h−1.
Keywords: Atomic absorption spectrometry; Flow injection; Continuous precipitation–dissolution; Iodine; Milk products;

Comparison of extracting solutions for elemental fractionation in airborne particulate matter by S. Canepari; M.L. Astolfi; S. Moretti; R. Curini (834-844).
It is here described the comparison of extraction efficiency of some solutions (acetate buffer, deionized water, diluted HNO3 and EDTA) frequently adopted in literature for evaluating the elemental solubility in airborne particulate matter. This comparison was performed considering the distribution of As, Ba, Ca, Cd, Co, Cr, Cu, Fe, Mg, Mn, Na, Ni, Pb, S, Si, Sb, Sn, Sr, Ti, V, Zn between the extractable and mineralized residual fractions on the NIST 1648 certified material, PM10 real samples and size-segregated samples, collected by a 13-stage impactor.The extracting solutions were evaluated by comparing extractive efficiencies and robustness towards some factors, such as acidity and concentration of complexing species, that have great environmental variability and that could be able to modify the extractive efficiency.Furthermore, extraction methods application to size-segregated samples allowed estimating the selectivity of extracting solutions towards dimensionally characterized emission sources, as dusts originated from abrasion and road dust re-suspension.On the basis of the obtained results, it was possible to define the main advantages and disadvantages resulting from the use of different extracting solutions, necessary to make possible the comparison of environmental studies carried out in different extractive conditions and to start up a proper study for harmonizing extracting procedures.
Keywords: Elemental solubility; Airborne particulate matter; Efficiency of extracting solutions; Selectivity of source tracers; Size-segregated samples;

An interference-free naked-eye recognition of Al3+ at its micromolar level has been done in 5% aqueous DMSO solution employing a Schiff base 5-[(2-hydroxy-5-nitro-benzylidene)-amino]-1H-pyrimidine-2,4-dione (receptor 1) which is an intramolecular charge transfer (ICT) probe. The pyrimidine and nitrophenyl groups serve as electron rich (donor) and deficient (acceptor) pockets in receptor 1 exhibiting a broad ICT band at 434 nm (olive green). The concomitant additions of Al3+ as its chloride salt to the 5 × 10−5  M aqueous DMSO solution of the receptor 1 lead hypsochromic shifting of its ICT band to 395 nm (colorless). The same ICT band undergoes a marginal bathochromic shifting (6 nm) along with a hyperchromic shift on separate additions of a basic anion like F, CH3COO and H2PO4 to the receptor 1 and faced almost similar fate on concomitant additions of Al3+ as mentioned above.
Keywords: Pyrimidine; 5-Aminouracil; Schiff base; Aluminium; Colorimetric receptor;

The paper presents an approach to use Partial Least Squares Discriminant Analysis (PLS-DA) on X-ray powder diffractometry (XRPD) dataset to build a model which recognizes a presence (or absence) of particular drug substance (acetaminophen) in unknown mixture (OTC tablet). The dataset consisted of 33 XRPD signals, measured for 12 pure substances and 21 tablets containing them in different quantitative and qualitative ratios, along with unknown excipients. The model was built with an external validation dataset chosen by Kennard–Stone algorithm. The RMSECV value was equal to 0.3461 (87.8% of explained variance) and external predictive error (RMSEP) was equal to 0.3123 (86.2% of explained variance). The result suggests that small but properly prepared training datasets give ability to construct well-working discriminant models on XRPD signals.
Keywords: Acetaminophen; Pharmaceuticals; X-ray powder diffractometry (XRPD); Chemometrics; Partial least squares;

Application of hollow fiber-based liquid-phase microextraction (HF-LPME) for the determination of acidic pharmaceuticals in wastewaters by María Ramos Payán; Miguel Ángel Bello López; Rut Fernández-Torres; Manuel Callejón Mochón; José Luis Gómez Ariza (854-858).
The presence of pharmaceuticals in the environment is a very important problem that requires analytical solutions. The wide variety of matrices and, usually, the low pharmaceuticals levels in the environmental samples requires high sensitive and selective analytical procedures. Wastewaters are one of the more important sources of environmental pollutants but they are very complex matrices that need clean-up procedures prior the analysis. Hollow fiber-based liquid-phase microextraction (HF-LPME) is a relatively new technique used in analytical chemistry for sample pre-treatment that offers high selectivity and sensitivity compared to most traditional extraction techniques. The low organic solvent consumption derived from the use of HF-LPME is according to the current trends to a “Green Chemistry”, and Analytical Chemistry should follow these environmental good practices. This paper describes an extraction method using a polypropylene membrane supporting dihexyl ether (three-phase hollow fiber-based liquid-phase microextraction (HF-LPME)) for the direct analysis of three pharmaceuticals (salicylic acid (SAC), ibuprofen (IBU) and diclofenac (DIC)) in raw and treated wastewaters followed by a HPLC/MS-MS determination using a highly packed Pursuit® XRs Ultra 2.8 μm C18 column that allows high resolution using low flow-rates and, simultaneously, short retention times. Detection limits were 20, 100 and 300 ng L−1 for salicylic acid, diclofenac and ibuprofen, respectively.
Keywords: Hollow fiber liquid-phase microextraction; HF-LPME; Wastewater; Salicylic acid; Diclofenac; Ibuprofen; HPLC–MS determination;

Ruthenium complexes containing 4,7-diphenyl-1,10-phenanthroline disulfonate (bathophenanthroline disulfonate; BPS) ligands, Ru(BPS)3 4−, Ru(BPS)2(bipy)2− and Ru(BPS)(bipy)2, were compared to tris(2,2′-bipyridine)ruthenium(II) (Ru(bipy)3 2+), including examination of the wavelengths of maximum absorption and corrected emission intensity, photoluminescence quantum yield, stability of their oxidised ruthenium(III) form, and relative chemiluminescence intensities and signal-to-blank ratios with cerium(IV) sulfate and six analytes (codeine, morphine cocaine, potassium oxalate, furosemide and hydrochlorothiazide) in acidic aqueous solution. The presence of BPS ligands in the complex increased the photoluminescence quantum yield, but decreased the stability of the oxidised form of the reagent. In contrast to previous evidence showing much greater electrochemiluminescence intensities using Ru(BPS)2(bipy)2− and Ru(BPS)(bipy)2, these complexes did not provide superior chemiluminescence signals than their homoleptic analogues.
Keywords: Chemiluminescence detection; Ruthenium complexes; Flow injection analysis;

by John Pickup (863).