Check out our New Publishers' Select for Free Articles

Analytical and Bioanalytical Chemistry (v.386, #6)

The curse of having a bad supervisor by John Fetzer (pp. 1577-1578).

is the author or co-author of over 130 research articles, reviews, and book chapters. He is a member of the International Advisory Board of Analytical and Bioanalytical Chemistry. Dr Fetzer worked for over 20 years as an analytical chemist for the Chevron Corporation and now runs his own consulting company, Fetzpahs Consulting, in Hercules, CA, USA. His book Career Management for Chemists—A Guide to Success in a Chemistry Career, was published by Springer.

Anna Tramontano: Protein structure prediction. Concepts and applications by Konrad Büssow (pp. 1579-1580).

Rohit Bhargava and Ira W. Levin (Eds): Spectrochemical analysis using infrared multichannel detectors by Reiner Salzer (pp. 1581-1582).

J.-L. Reymond (Ed.): Enzyme assays. High-throughput screening, genetic selection and fingerprinting by Will Bloch (pp. 1583-1584).

Katherine A. Bakeev (Ed.): Process Analytical Technology by Stephan Küppers (pp. 1585-1586).

Labeling the human skeleton with ⁴¹Ca to assess changes in bone calcium metabolism by E. Denk; D. Hillegonds; J. Vogel; A. Synal; C. Geppert; K. Wendt; K. Fattinger; C. Hennessy; M. Berglund; R. F. Hurrell; T. Walczyk (pp. 1587-1602).

Bone research is limited by the methods available for detecting changes in bone metabolism. While dual X-ray absorptiometry is rather insensitive, biochemical markers are subject to significant intra-individual variation. In the study presented here, we evaluated the isotopic labeling of bone using ⁴¹Ca, a long-lived radiotracer, as an alternative approach. After successful labeling of the skeleton, changes in the systematics of urinary ⁴¹Ca excretion are expected to directly reflect changes in bone Ca metabolism. A minute amount of ⁴¹Ca (100 nCi) was administered orally to 22 postmenopausal women. Kinetics of tracer excretion were assessed by monitoring changes in urinary ⁴¹Ca/⁴⁰Ca isotope ratios up to 700 days post-dosing using accelerator mass spectrometry and resonance ionization mass spectrometry. Isotopic labeling of the skeleton was evaluated by two different approaches: (i) urinary ⁴¹Ca data were fitted to an established function consisting of an exponential term and a power law term for each individual; (ii) ⁴¹Ca data were analyzed by population pharmacokinetic (NONMEM) analysis to identify a compartmental model that describes urinary ⁴¹Ca tracer kinetics. A linear three-compartment model with a central compartment and two sequential peripheral compartments was found to best fit the ⁴¹Ca data. Fits based on the use of the combined exponential/power law function describing urinary tracer excretion showed substantially higher deviations between predicted and measured values than fits based on the compartmental modeling approach. By establishing the urinary ⁴¹Ca excretion pattern using data points up to day 500 and extrapolating these curves up to day 700, it was found that the calculated ⁴¹Ca/⁴⁰Ca isotope ratios in urine were significantly lower than the observed ⁴¹Ca/⁴⁰Ca isotope ratios for both techniques. Compartmental analysis can overcome this limitation. By identifying relative changes in transfer rates between compartments in response to an intervention, inaccuracies in the underlying model cancel out. Changes in tracer distribution between compartments were modeled based on identified kinetic parameters. While changes in bone formation and resorption can, in principle, be assessed by monitoring urinary ⁴¹Ca excretion over the first few weeks post-dosing, assessment of an intervention effect is more reliable ∼150 days post-dosing when excreted tracer originates mainly from bone.

Keywords: ⁴¹Ca; AMS; RIMS; Osteoporosis; Kinetic analysis; Compartmental modeling

Differential proteome analysis of colon carcinoma cell line SW480 after reconstitution of the tumour suppressor Smad4 by Kai Stühler; Katharina Köper; Kathy Pfeiffer; Andreas Tagariello; Manfred Souquet; Irmgard Schwarte-Waldhoff; Stephan A. Hahn; Wolff Schmiegel; Helmut E. Meyer (pp. 1603-1612).

The tumour suppressor gene Smad4 is frequently inactivated in gastrointestinal carcinomas. Smad4 plays a pivotal role in transducing signals of the transforming growth factor-β (TGF-β) superfamily of proteins. Inactivation of Smad4 seems to occur late during tumour progression when tumours acquire invasive and metastatic properties. Identification of proteins directly or indirectly regulated by Smad4 would, therefore, ease the future design of new diagnostic and therapeutic strategies for gastrointestinal carcinoma. We have used human colon carcinoma cell line SW480 stably transfected with Smad4 as an in-vitro model system to identify Smad4-regulated proteins by applying two-dimensional gel electrophoresis (2DE) then MALDI-PMF/PFF-MS. We identified a total of 47 protein species with a Smad4-dependent expression. From the functions of the candidate proteins we obtained new insights into Smad4’s participation in processes, for example apoptosis, differentiation, and proliferation.

Keywords: Differential proteome analysis; Two-dimensional gel electrophoresis; DPC4; Smad4 ; Colorectal cancer; Tumour suppressor

Optimization of DNA-tagged liposomes for use in microtiter plate analyses by Katie A. Edwards; Antje J. Baeumner (pp. 1613-1623).

Dye-encapsulating unilamellar DNA oligonucleotide-tagged liposomes were prepared and characterized for use as signal-enhancing reagents in a microtiter plate sandwich-hybridization analyses of single-stranded RNA or DNA sequences. The liposomes were synthesized using the reversed-phase evaporation method and tagged with DNA oligonucleotides by adding cholesteryl-modified DNA reporter probes to the initial lipid mixture. Liposomes were prepared using probe coverages of 0.0013–0.103 mol% of the total lipid input, several hydrophobic and poly(ethylene glycol)-based spacers between the cholesteryl anchor and the probe, and liposome diameters ranging from 200 nm to 335 nm. Their signal enhancement functionality was compared by using them in microtiter plate sandwich-hybridization assays for the detection of single-stranded DNA sequences. In these assays, an optimal reporter probe concentration of 0.103 mol%, a liposome diameter of 274 nm, and a phospholipid concentration of 0.3 mM were found. The length between the cholesteryl anchor and the probe was optimal when a spacer composed of TEG+(CH₂O)₃ was used. Under optimal conditions, a detection limit of 0.5 nM for a truncated synthetic DNA sequence was found with a coefficient of variation of 4.4%. A 500-fold lower limit of detection using fluorescence was found using lysed dye-encapsulating liposomes versus a single fluorescein-labeled probe. Finally, when this method was applied to the detection of atxA RNA extracted from E.coli SG12036-pIu121 and amplified using NASBA, a minimum extracted concentration of RNA of 1.1×10⁻⁷ μg/μL was found.

Keywords: Liposomes; DNA; Sandwich-hybridization; Conjugation; Fluorescence; Biosensor

Development of RNR3- and RAD54-GUS reporters for testing genotoxicity in Saccharomyces cerevisiae by Susanna Boronat; Benjamin Piña (pp. 1625-1632).

S. cerevisiae RNR3 and RAD54 gene transcription becomes strongly activated upon DNA damage. This property was used to construct yeast strains in which DNA damage can be monitored by a very sensitive fluorogenic assay in a convenient 96-well microtiter plate format. These strains carried stably integrated fusions of RNR3 or RAD54 promoters to the E. coli β-glucuronidase GUS gene. GUS activity was measured by fluorogenic detection, a method that greatly increases the precision and sensitivity of the assay. Detection levels were similar to those of real-time quantitative PCR methods and close to the limits of biological response. The two reporters differed in terms of fold-induction, activation kinetics, sensitivity and specificity upon exposure to a variety of genotoxic compounds. While RNR3-GUS showed the fastest response, RAD54-GUS showed the highest sensitivity: similar to previous reported sensitivities for bacterial and eukaryotic genotoxic detection systems. These reporter strains may complement current genotoxicity tests, but they also have the advantages of higher flexibility, requirement for shorter incubation times, and the capability of being fully automated. In addition, the intrinsic features of the system facilitate its easy improvement by genetic manipulating the yeast strain or by introducing mammalian metabolizing enzymes.

Keywords: DNA repair; Mutagenesis; Biosensors; Fluorogenic reporters; Cytotoxicity; Yeast bioassays

In vitro nonenzymatic glycation of DNA nucleobases: an evaluation of advanced glycation end products under alkaline pH by Udayan Dutta; Menashi A. Cohenford; Madhumita Guha; Joel A. Dain (pp. 1633-1640).

The advanced glycation end products (AGEs) of DNA nucleobases have received little attention, perhaps due to the fact that adenine, guanine, cytosine and thymine do not dissolve under mild pH conditions. To maintain nucleobases in solution, alkaline pH conditions are typically required. The objectives of this investigation were twofold: to study the susceptibility of DNA nucleobases to nonenzymatic attack by different sugars, and to evaluate the factors that influence the formation of nucleobase AGEs at pH 12, i.e., in an alkaline environment that promotes the aldo–keto isomerization and epimerization of sugars. Varying concentrations of adenine, guanine, thymine and cytosine were incubated over time with constant concentrations of D-glucose, D-galactose or D/L-glyceraldehyde under different conditions of temperature and ionic strength. Incubation of the nucleobases with the sugars resulted in a heterogeneous assembly of AGEs whose formation was monitored by UV/fluorescence spectroscopy. Capillary electrophoresis and HPLC were used to resolve the AGEs of the DNA adducts and provided a powerful tool for following the extent of glycation in each of the DNA nucleobases. Mass spectrometry studies of DNA adducts of guanine established that glycation at pH 12 proceeded through an Amadori intermediate.

Keywords: Advanced glycation end products; AGEs; Nucleobase glycation

Tagging of avidin immobilized beads with biotinylated YAG:Ce³⁺ nanocrystal phosphor by Ryo Asakura; Tetsuhiko Isobe; Kiyoshi Kurokawa; Hideki Aizawa; Michio Ohkubo (pp. 1641-1647).

YAG:Ce³⁺ nanoparticles 9.5 ± 1.2 nm in diameter have been synthesized from aluminium isopropoxide and acetates of yttrium and cerium in 1,4-butanediol (1,4-BD) by autoclave treatment at 300 °C for 2 h. After replacing 1,4-BD by ultrapure water, NH₂ groups were introduced on the surface of YAG:Ce³⁺ nanoparticles by addition of 3-aminopropyltrimethoxysilane then biotinylation with sulfo-NHS-LC-biotin. We demonstrated that avidin immobilized beads are tagged by biotinylated YAG:Ce³⁺ nanoparticles by the selective avidin-biotin interaction, furnishing a green fluorescent image on excitation with blue light. This result indicates that YAG:Ce³⁺ nanoparticle phosphors have much potential in biological applications.

Keywords: YAG:Ce³⁺ ; Nanoparticle; 3-aminopropyltrimethoxysilane; Avidin-biotin interaction; Fluorescent image; Biological application

Development of an efficient amine-functionalized glass platform by additional silanization treatment with alkylsilane by Nagendra Kumar Kamisetty; Seung Pil Pack; Mitsuru Nonogawa; Kamakshaiah Charyulu Devarayapalli; Tsutomu Kodaki; Keisuke Makino (pp. 1649-1655).

Aminosilane-treated molecular layers on glass surfaces are frequently used as functional platforms for biosensor preparation. All the amino groups present on the surface are not available in reactive forms, because surface amino groups interact with remaining unreacted surface silanol groups. Such nonspecific interactions might reduce the efficiency of chemical immobilization of biomolecules such as DNA, enzymes, antibodies, etc., in biosensor fabrication. To improve immobilization efficiency we have used additional surface silanization with alkylsilane (capping) to convert the remaining silanol groups into Si–O–Si linkages, thereby liberating the amino groups from nonspecific interaction with the silanol groups. We prepared different types of capped amine surface and evaluated the effect of capping on immobilization efficiency by investigating the fluorescence intensity of Cy3-NHS (N-hydroxysuccinimide) dye that reacted with amino groups. The results indicate that most of the capped amine surfaces resulted in enhanced efficiency of immobilization of Cy3-NHS compared with the untreated control amine surface. We found a trend that trialkoxysilanes had greater capping effects on immobilization efficiency than monoalkoxysilanes. It was also found that the aliphatic chain of alkylsilane, which does not participate in the capping of the silanol, had an important function in enhancing immobilization efficiency. These results would be useful for preparation of an amine-modified surface platform, with enhanced immobilization efficiency, which is essential for developing many kinds of biosensors on a silica matrix. Enhancement of amine funtionality by capping with alkylsilane

Keywords: Aminosilane; Glass surface modification; Capping

Development of a competitive ELISA for the quantification of F5 conjugate in HER2-targeted STEALTH immunoliposome doxorubicin in plasma samples by Ran Hu; Rebecca Davis; Yongjin Yao; Yaodong Xu (pp. 1657-1664).

HER2 (human epidermal growth factor receptor 2, erbB2, or neu) is overexpressed by a large number of tumor types and has been identified as an important target for cancer therapy. F5 is a single-chain human antibody fragment that recognizes HER2 receptor and is covalently conjugated to PEGylated lipid to form F5 conjugate (F5CG) in the product HER2 targeted STEALTH immunoliposome doxorubicin. Here we described the method development of a competitive enzyme-linked immunosorbent assay (ELISA) for the determination of total concentration of F5 conjugate in plasma samples. The method involved the biotinylation of F5CG, detergent treatment of plasma sample to solubilize F5CG into monomeric form, and competitive ELISA for solubilized F5CG competitively binding to anti-F5CG antibody with biotinylated F5CG for the determination of total F5CG in plasma. The detection range of this method was from 0.2 ng/mL to 125 ng/mL for F5CG in plasma. The lower limit of quantification (LLOQ) was 0.2 ng/mL. This method was established and used for the measurement of F5CG concentration to provide information about F5CG circulation after the administration of immunoliposome in preclinical studies.

Keywords: Competitive ELISA; Assay development; F5 conjugate (F5CG); Targeted immunoliposomal formulation; HER2 receptor

Characterization of the coupling of quantum dots and immunoglobulin antibodies by Xiao-Feng Hua; Tian-Cai Liu; Yuan-Cheng Cao; Bo Liu; Hai-Qiao Wang; Jian-Hao Wang; Zhen-Li Huang; Yuan-Di Zhao (pp. 1665-1671).

Water-soluble quantum dots (QDs) were used to label goat anti-human immunoglobulin antibodies (Abs), and the labeling process was characterized by column purification. The QDs obtained in organic solvent were modified with mercaptoacetic acid (MAA) and became water-soluble. These water-soluble QDs were linked to the antibodies using the coupling reagents ethyl-3-(dimethyl aminopropyl) carbodiimide (EDC) and N-hydroxysuccinimide (NHS). The linking process was shown to be effective by ultra-filter centrifugation and column purification. After comparing the quantities of Abs and water-soluble QDs involved in the linking reaction via column purification, it was found that a molar Abs:QD ratio of >1.2 resulted in most of the water-soluble QDs becoming covalently linked to the Abs. The circular dichroism (CD) spectra of Abs and QD–Ab conjugates were very similar to each other, indicating that the secondary structure of Abs remained largely intact after the conjugation. Finally, antigen (Ag)–antibody (Ab) recognition reactions perfomed on the surface of a glass slide showed that the conjugate retained the activity of Abs. This work lends support to the idea of linking biomolecules to QDs, and thus should aid the application of QDs to the life sciences. Figure Firstly in this work, the conjugates of QDs-Ab were separated from EDC&NHS in the column of Sephadex G-100(left up). Then the bioactivity of QDs-Ab was analyzed in the immunoassay (right) and the immunofluorescent signals were detected (left bottom) finally

Keywords: Fluorescent labeling; Quantum dots; Bioconjugate; Antigen; Antibody

Application of double-spike isotope dilution for the accurate determination of Cr(III), Cr(VI) and total Cr in yeast by Lu Yang; Elena Ciceri; Zoltán Mester; Ralph E. Sturgeon (pp. 1673-1680).

A method is presented for the simultaneous determination of Cr(III) and Cr(VI) in yeast using species-specific double-spike isotope dilution (SSDSID) with anion-exchange liquid chromatography (LC) separation and sector field inductively coupled plasma mass spectrometric (SF-ICP-MS) detection. Total Cr is quantitated using ID SF-ICP-MS. Samples were digested on a hot plate at 95±2 °C for 6 h in an alkaline solution of 0.5 M NaOH and 0.28 M Na₂CO₃ for the determination of Cr(III) and Cr(VI), whereas microwave-assisted decomposition with HNO₃ and H₂O₂ was used for the determination of total Cr. Concentrations of 2,014±16, 1,952±103 and 76±48 mg kg⁻¹ (one standard deviation, n=4, 3, 3), respectively were obtained for total Cr, Cr(III) and Cr(VI) in the yeast sample. Significant oxidation of Cr(III) to Cr(VI) (24.2±7.6% Cr(III) oxidized, n=3) and reduction of Cr(VI) to Cr(III) (37.6±6.5% Cr(VI) reduced, n=3 ) occurred during alkaline extraction and subsequent chromatographic separation at pH 7. Despite this significant bidirectional redox transformation, quantitative recoveries for both Cr(III) and Cr(VI) were achieved using the SSDSID method. In addition, mass balance between total Cr and the sum of Cr(III) and Cr(VI) concentrations was achieved. Method detection limits of 0.3, 2 and 30 mg kg⁻¹ were obtained for total Cr, Cr(VI) and Cr(III), respectively, based on a 0.2-g sub-sample.

Keywords: Cr(III); Cr(VI); Double-spike isotope dilution; Yeast; ICP-MS

Observation of salt-induced β-lactoglobulin aggregation using sedimentation field-flow fractionation by Sudarat Saeseaw; Juwadee Shiowatana; Atitaya Siripinyanond (pp. 1681-1688).

Sedimentation field-flow fractionation (SdFFF) was applied in order to characterize particle sizes of β-lactoglobulin aggregates induced by Ca²⁺ or Zn²⁺. Aggregation induced by Zn²⁺ was faster than that induced by Ca²⁺. Effects of Zn²⁺ and β-lactoglobulin concentrations, as well as contact time, on the aggregation of β-lactoglobulin were examined. All factors exhibited a combined effect on the size of aggregates, whereby larger aggregates were obtained at increased concentrations of Zn²⁺ and β-lactoglobulin. At fixed concentrations of 2% (w/v) β-lactoglobulin and 10 mM Zn²⁺, the particle size of the aggregates increased from 0.19 μm (at 15 min) to 0.38 μm (at 2880 min). Further, a hyphenated technique of SdFFF and inductively coupled plasma–optical emission spectrometry (ICP–OES) was used to examine whether intermolecular ionic bridges take part in salt-induced β-lactoglobulin aggregation. With SdFFF–ICP–OES, protein–cation–protein cross-linkages were observed for β-lactoglobulin aggregation induced by Zn²⁺, but not for that induced by Ca²⁺.

Keywords: Lactoglobulin; Aggregation; Field-flow fractionation; Inductively coupled plasma–optical emission spectrometry; Calcium and zinc ions

Determination of the titanium content of human transferrin by inductively-coupled plasma-atomic-emission spectroscopy by Aracelis Cardona; Enrique Meléndez (pp. 1689-1693).

We report a simple method that combines dialysis, as a purification method, with the multielement capability of ICP to determine the titanium-to-transferrin mole ratio at physiological pH, under buffer conditions. The method, by means of which titanium and transferrin are determined simultaneously, enabled us to assess the binding capacities of different titanocene complexes. Figure Titanocene dichloride

Keywords: Titanocene dichloride; ICP-AES; Transferrin; Antitumor

Phototoxicity testing by online irradiation and HPLC by Sven Schröder; J. P. Surmann (pp. 1695-1700).

A high-performance liquid chromatography (HPLC) system was developed for the determination of drug photostability and phototoxicity based on an automated column-switching system with aqueous online UV-A irradiation and hyphenated organic separation of the drug and its photoproducts. The photoreactor is built with an poly(ethylene-co-tetrafluoroethylene) (ETFE) reaction coil knitted around a UV-A light source. The chromatographic separation was performed with two special C₁₈ columns, which are also suitable for using with pure water as eluent. Degradation of chlorpromazine (CPZ) by ultraviolet light was investigated at pH 7 and pH 3. Furthermore chlorpromazine was irradiated in the presence of guanosine-5-monophosphate (GMP) in pH 7 buffered solution, leading to a new photoproduct. In the pH 3 irradiation studies of CPZ and GMP, no reaction was detected between the molecules.

Keywords: Drug monitoring/screening; Pharmaceuticals; Nucleic acids (DNA/RNA); HPLC; Photochemistry; Phototoxicity

Simulation of the detoxification of paracetamol using on-line electrochemistry/liquid chromatography/mass spectrometry by Wiebke Lohmann; Uwe Karst (pp. 1701-1708).

On-line electrochemistry/liquid chromatography/mass spectrometry was used to simulate the detoxification mechanism of paracetamol in the body. In an electrochemical flow-through cell, paracetamol was oxidized at a porous glassy carbon working electrode at a potential of 600 mV vs. Pd/H₂ with formation of a quinoneimine intermediate. The quinoneimine further reacted with glutathione and/or N-acetylcysteine to form isomeric adducts via the thiol function. The adducts were characterized on-line by liquid chromatography/mass spectrometry. These reactions are similar to those occurring between paracetamol and glutathione under catalysis by cytochrome P450 enzymes in the body.

Keywords: Paracetamol (acetaminophen, p-acetamidophenol); Detoxification; N-Acetylcysteine; Glutathione; Electrochemistry; Liquid chromatography/mass spectrometry

Site-specific acid–base properties of pholcodine and related compounds by Z. Kovács; S. Hosztafi; B. Noszál (pp. 1709-1716).

The acid–base properties of pholcodine, a cough-depressant agent, and related compounds including metabolites were studied by ¹H NMR-pH titrations, and are characterised in terms of macroscopic and microscopic protonation constants. New N-methylated derivatives were also synthesized in order to quantitate site- and nucleus-specific protonation shifts and to unravel microscopic acid–base equilibria. The piperidine nitrogen was found to be 38 and 400 times more basic than its morpholine counterpart in pholcodine and norpholcodine, respectively. The protonation data show that the molecule of pholcodine bears an average of positive charge of 1.07 at physiological pH, preventing it from entering the central nervous system, a plausible reason for its lack of analgesic or addictive properties. The protonation constants of pholcodine and its derivatives are interpreted by comparing with related molecules of pharmaceutical interest. The pH-dependent relative concentrations of the variously protonated forms of pholcodine and morphine are depicted in distribution diagrams.

Keywords: Pholcodine; Metabolites; Acid–base equilibria; Microspeciation; Protonation

Determination of parathion in biological fluids by means of direct solid-phase microextraction by E. Gallardo; M. Barroso; C. Margalho; A. Cruz; D. N. Vieira; M. López-Rivadulla (pp. 1717-1726).

A new and simple procedure for the determination of parathion in human whole blood and urine using direct immersion (DI) solid-phase microextraction (SPME) and gas chromatography/mass spectrometry (GC/MS) is presented. This technique was developed using only 100 μL of sample, and ethion was used as internal standard (IS). A 65-μm Carbowax/divinylbenzene (CW/DVB) SPME fibre was selected for sampling, and the main parameters affecting the SPME process such as extraction temperature, adsorption and desorption time, salt addition, agitation and pH effect were optimized to enhance the sensitivity of the method. This optimization was also performed to allow the qualitative determination of parathion’s main metabolite, paraoxon, in blood. The limits of detection and quantitation for parathion were 3 and 10 ng/mL for urine and 25 and 50 ng/mL for blood, respectively. For paraoxon, the limit of detection was 50 ng/mL in blood. The method showed linearity between the LOQ and 50 μg/mL for both matrices, with correlation coefficients ranging from 0.9954 to 0.9999. Precision and accuracy were in conformity with the criteria normally accepted in bioanalytical method validation. The mean absolute recoveries were 35.1% for urine and 6.7% for blood. Other parameters such as dilution of sample and stability were also validated. Its simplicity and the fact that only 100 μL of sample is required to accomplish the analysis make this method useful in forensic toxicology laboratories to determine this compound in intoxications, and it can be considered an alternative to other methods normally used for the determination of this compound in biological media.

Keywords: Direct immersion solid-phase microextraction; Parathion; Whole blood; Urine

Determination of ruscogenin in crude Chinese medicines and biological samples by immunoassay by Nan Liu; Xingbing Wen; Jihua Liu; Ming Liang; Huajin Zeng; Yining Lin; Boyang Yu (pp. 1727-1733).

Ruscogenin is a major bioactive steroidal aglycone found in the Chinese medicine, Ophiopogon japonicus. We have developed a quantitative determination of ruscogenin with an indirect enzyme-linked immunosorbent (ELISA) assay using polyclonal antibodies against ruscogenin conjugated with bovine serum albumin. This assay was highly sensitive, and it had considerably less cross-reactivity to diosgenin and sarsasapogenin, but high cross-reactivity to ruscogenin glycosides. The assay was successfully used for the measurement of ruscogenin concentrations in crude Chinese medicines and in biological samples from a pharmacokinetics study of ruscogenin.

Keywords: Ruscogenin; Immunoassay; Pharmacokinetics; Biological sample; Crude Chinese medicine

Development of a monoclonal antibody-based enzyme-linked immunosorbent assay for the analysis of glycyrrhizic acid by Jing Zhao; Gang Li; Bao-min Wang; Wei Liu; Tie-gui Nan; Zhi-xi Zhai; Zhao-hu Li; Qing X. Li (pp. 1735-1740).

Glycyrrhizic acid (GL) is a major active compound of licorice. The specific monoclonal antibody (MAb) (designated as 8F₈A₈H₄2H₇) against GL was produced with the immunogen GL–BSA conjugate. The dissociation constant (K _d) value of the MAb was approximately 9.96×10⁻¹⁰ M. The cross reactivity of the MAb with glycyrrhetic acid was approximately 2.6%. The conventional indirect competitive enzyme-linked immunosorbent assay (icELISA) and simplified icELISA adapted with a modified procedure were established using the MAb. The IC₅₀ value and the detect range by the conventional icELISA were 1.1 ng mL⁻¹ and 0.2–5.1 ng mL⁻¹, respectively. The IC₅₀ value and the detect range by the simplified icELISA were 5.3 ng mL⁻¹ and 1.2–23.8 ng mL⁻¹, respectively. The two icELISA formats were used to analyze GL contents in the roots of wild licorice and different parts of cultivated licorice (Glycyrrhiza uralensis Fisch). The results obtained with the two icELISAs agreed well with those of the HPLC analysis. The correlation coefficient was more than 0.98 between HPLC and the two icELISAs. The two icELISAs were shown to be appropriate, simple, and effective for the quality control of raw licorice root materials.

Keywords: Glycyrrhizic acid; Monoclonal antibody; ELISA; Licorice

Fluorescence determination of metoprolol in human plasma by trilinear decomposition-based calibration techniques by Yan Zhang; Hai-Long Wu; A-Lin Xia; Shao-Hua Zhu; Qing-Juan Han; Ru-Qin Yu (pp. 1741-1748).

In this study a new spectrofluorimetric method for the direct determination of metoprolol in human plasma is presented and discussed. It is based on the use of fluorescence excitation–emission matrices (EEMs) and second-order calibration performed with parallel factor analysis (PARAFAC) or alternating trilinear decomposition (ATLD). This methodology enables accurate and reliable discrimination of the analyte signal, even in the presence of unknown and uncalibrated fluorescent component(s), which is often referred to as the second-order advantage. No separation or sample pretreatment steps were required. Satisfactory results were obtained. Metoprolol recoveries in plasma were determined as 87±2% and 90±4% with PARAFAC and ATLD, respectively. All RSD values of intra- and interday assays were below 5%. Figure A three-dimensional plot of EEMs for a plasma sample and metoprolol solution

Keywords: Metoprolol; Plasma; PARAFAC; ATLD; Fluorescence excitation–emission matrices

Development of a supercritical fluid extraction–gas chromatography–mass spectrometry method for the identification of highly polar compounds in secondary organic aerosols formed from biogenic hydrocarbons in smog chamber experiments by L. Chiappini; E. Perraudin; R. Durand-Jolibois; J. F. Doussin (pp. 1749-1759).

A new one-step method for the analysis of highly polar components of secondary organic aerosols (SOA) has been developed. This method should lead to a better understanding of SOA formation and evolution since it enables the compounds responsible for SOA formation to be identified. Since it is based on supercritical fluid extraction coupled to gas chromatography–mass spectrometry, it minimizes the analysis time and significantly enhances sensitivity, which makes it suitable for trace-level compounds, which are constituents of SOA. One of the key features of this method is the in situ derivatisation step: an online silylation allowing the measurement of highly polar, polyfunctional compounds, which is a prerequisite for the elucidation of chemical mechanisms. This paper presents the development of this analytical method and highlights its ability to address this major atmospheric issue through the analysis of SOA formed from the ozonolysis of a biogenic hydrocarbon (sabinene). Ozonolysis of sabinene was performed in a 6 m³ Teflon chamber. The aerosol components were derivatised in situ. More than thirty products, such as sabinaketone, sabinic acid and other multifunctional compounds including dicarboxylic acids and oxoacids, were measured. Nine of them were identified and quantified. The sensitivity and the linearity (0.91 < R < 0.98) of the method were both good and detection limits ranged from 1.2 to 6.4 ng for the investigated compounds.

Keywords: Secondary organic aerosol; Chemical composition; Supercritical fluid extraction; Sabinene; Highly polar compounds

Microstructure and chemical composition of giant avian eggshells by Yannicke Dauphin; Jean-Pierre Cuif; Murielle Salomé; Jean Susini; C. Terry Williams (pp. 1761-1771).

The microstructure and composition of the layers of two giant avian eggshells were investigated using a combination of scanning electron microscopy, electron probe microanalyses, and X-ray absorption near-edge structure spectroscopy (XANES). The two species have some similarities and differences in their microstructure and composition; the composition is not homogeneous throughout the eggshell thickness. XANES studies show that sulfur is associated with amino acids in the inner organic membranes, whereas in the mineralised layers the sulfur is mainly associated with sulfated polysaccharides. These results are similar to those obtained on chicken eggshells, and confirm the active role of sulfated acidic polysaccharides in biomineralisation processes of carbonate skeletons.

Keywords: Biomineral; Eggshell; Aves; Sulfur; EPMA; XANES

A rapid analytical method for predicting the oxygen demand of wastewater by Shoshana Fogelman; Huijun Zhao; Michael Blumenstein (pp. 1773-1779).

In this study, an investigation was undertaken to determine whether the predictive accuracy of an indirect, multiwavelength spectroscopic technique for rapidly determining oxygen demand (OD) values is affected by the use of unfiltered and turbid samples, as well as by the use of absorbance values measured below 200 nm. The rapid OD technique was developed that uses UV–Vis spectroscopy and artificial neural networks (ANNs) to indirectly determine chemical oxygen demand (COD) levels. It was found that the most accurate results were obtained when a spectral range of 190–350 nm was provided as data input to the ANN, and when using unfiltered samples below a turbidity range of 150 NTU. This is because high correlations of above 0.90 were obtained with the data using the standard COD method. This indicates that samples can be measured directly without the additional need for preprocessing by filtering. Samples with turbidity values higher than 150 NTU were found to produce poor correlations with the standard COD method, which made them unsuitable for accurate, real-time, on-line monitoring of OD levels.

Keywords: Chemical oxygen demand; Wastewater monitoring; UV–Vis spectrometry; Artificial neural networks; Multicomponent analysis

Comparison of the efficiencies of different types of adsorbents at trapping currently used pesticides in the gaseous phase using the technique of high-volume sampling by Rowan Dobson; Anne Scheyer; Anne Laure Rizet; Philippe Mirabel; Maurice Millet (pp. 1781-1789).

Atmospheric samples were collected in an urban area (Strasbourg centre) in spring/summer 2004, in order to determine the concentrations of different pesticides in the gaseous and particulate phases and to compare the efficiencies of different adsorbents at trapping the gaseous phase. Two high-volume samplers were placed next to each other in the botanical garden in the centre of Strasbourg. Air sampling was carried out using a glass fibre filter and different adsorbents for 48 hrs. The following adsorbents and combinations of adsorbents were compared: XAD-2 with PUF, XAD4 with PUF, XAD-2 with a PUF-XAD2-PUF sandwich, PUF with a PUF-XAD4-PUF sandwich. In order of efficiency at trapping pesticides, the “sandwiches” are the most efficient, followed by XAD-2 and XAD-4 resins. However, although the “sandwiches” are slightly better at trapping than XAD-2, the use of XAD-2 is recommended for technical reasons. The PUFs are the least efficient at trapping. Among the 27 pesticides analysed, trifluralin, alachlor, metolachlor and captan were the most concentrated pesticides, followed by lindane, alpha-endosulfan and diflufenican. This result is in accordance with farming activity in the Alsace region, where the pesticides that are used on large crops (maize, cereals) are applied in the greatest quantities. Vineyards are another important form of agriculture in Alsace, but the quantities of pesticides applied in comparison to those used on large crops is very low, which explains the low detection of vineyard pesticides in air samples observed here. The concentrations are depend on the identities and properties of the pesticides analysed, but on the whole they remain rather low. It is important to perform measurements like these in the urban environment, as these compounds can be harmful to human health and the environment and so their concentrations need to be monitored.

Keywords: Pesticides; Urban areas; Solid adsorbents; Gaseous phase; Particulate phase

Characterization of volatile organic compounds and odors by in-vivo sampling of beef cattle rumen gas, by solid-phase microextraction, and gas chromatography–mass spectrometry–olfactometry by Lingshuang Cai; Jacek A. Koziel; Jeremiah Davis; Yin-Cheung Lo; Hongwei Xin (pp. 1791-1802).

Volatile organic compounds (VOCs) and odors in cattle rumen gas have been characterized by in-vivo headspace sampling by solid-phase microextraction (SPME) and analysis by gas chromatography–mass spectrometry–olfactometry (GC–MS–O). A novel device enabling headspace SPME (HS-SPME) sampling through a cannula was designed, refined, and used to collect rumen gas samples from steers. A Carboxen–polydimethylsiloxane (PDMS) fiber (85 μm) was used for SPME sampling. Fifty VOCs from ten chemical groups were identified in the rumen headspace. The VOCs identified had a wide range of molecular weight (MW) (34 to 184), boiling point (−63.3 to 292 °C), vapor pressure (1.05 × 10⁻⁵ to 1.17 × 10² Pa), and water solubility (0.66 to 1 × 10⁶ mg L⁻¹). Twenty-two of the compounds have a published odor detection thresholds (ODT) of less than 1 ppm. More than half of the compounds identified are reactive and have an estimated atmospheric lifetime of <24 h. The amounts of VFAs, sulfide compounds, phenolic compounds, and skatole, and the odor intensity of VFAs and sulfide compounds in the rumen gas were all higher after feeding than before feeding. These results indicate that rumen gases can be an important potential source of aerial emissions of reactive VOCs and odor. In-vivo sampling by SPME then GC–MS–O analysis can be a useful tool for qualitative characterization of rumen gases, digestion, and its relationship to odor and VOC formation. Figure Modified cannula for rumen gas sampling with SPME

Keywords: Rumen gas; Odor; In-vivo sampling; SPME; GC–MS–O

Kinetic speciation of nickel in mining and municipal effluents by Parthasarathi Chakraborty; Yamini Gopalapillai; John Murimboh; Ismail I. Fasfous; Chuni L. Chakrabarti (pp. 1803-1813).

This study presents the results of kinetic speciation of nickel in undiluted mining and municipal effluents and effluents diluted with receiving freshwaters from the surrounding environment. The dilution ratios used for the dilution of the effluents were arbitrarily chosen, but were representative of the prevailing mining practices. The purpose of the this dilution was to mimic dilution with natural waters that result from dilution of the mining and municipal effluents with receiving freshwaters, so that this study would reveal environmental realities that are of concern to the managers and regulators of water resources.Ligand exchange kinetics using the competing ligand exchange method (CLEM) was studied using two independent techniques: graphite furnace atomic absorption spectrometry (GFAAS) with Chelex 100 resin as the competing ligand, and adsorptive cathodic stripping voltammetry (AdCSV) with dimethylglyoxime (DMG) as the competing ligand to determine the percentage of Ni metal released from Ni(II)–DOC complexes and the rate of dissociation of Ni(II)–DOC complexes. Using a sample containing a mixture of 30% Copper Cliff Mine effluent, 40% Sudbury municipal effluent and 30% Vermillion River water, both techniques gave results showing that the dilution of the effluent samples increased the percentage of nickel released from Ni(II)–DOC complexes. This increase in the release of nickel from the Ni(II)–DOC complexes may be of concern to managers and regulators of water resources. Agreement between the results of these two techniques has enhanced the validity of the competing ligand exchange method used by both techniques.

Keywords: Nickel speciation; Mine effluent; Competing ligand exchange method; Release of nickel; Dissolved Organic Carbon; Adsorptive Cathodic Stripping Voltammetry; Graphite Furnace Atomic Absorption Spectrometry

Compositional and technological features of glazed pottery from Aosta Valley (Italy): a SEM–EDS investigation by Monica Gulmini; Lorenzo Appolonia; Patrizia Framarin; Piero Mirti (pp. 1815-1822).

Twelve finds from archaeological excavations carried out in the Aosta region (Italy) were studied by scanning electron microscopy coupled with energy-dispersive X-ray detection (SEM–EDS). The archaeological samples were shards of glazed pottery dating from the fourth to the seventh century AD. Analysis of ceramic bodies revealed a general homogeneity in composition among the studied samples and the use of a noncalcareous clay for their manufacture; however, two shards stand out due to their high iron contents. Glazes proved to be high-lead products with more than 70% PbO in all of the samples investigated but one. For the latter, a composition poorer in lead and richer in silicon, aluminium and iron was found. SEM observation of the contact region between body and glaze suggests that the vitreous coatings were mostly obtained by applying the glazing components onto the unfired clay body; moreover, a comparison between clay and glaze compositions suggests the use of a lead compound mixed with a silica-rich material, not a lead compound by itself.

Keywords: Lead glaze; Pottery technology; Scanning electron microscopy; Energy-dispersive X-ray analysis

Near-infrared reflectance spectroscopy as a fast and non-destructive tool to predict foliar organic constituents of several woody species by C. Petisco; B. García-Criado; S. Mediavilla; B. R. Vázquez de Aldana; I. Zabalgogeazcoa; A. García-Ciudad (pp. 1823-1833).

Near-infrared reflectance spectroscopy (NIRS) was used to estimate N, neutral detergent fibre (NDF), acid detergent fibre (ADF), lignin and cellulose contents in leaves of a heterogeneous group of 17 woody species from the Central Western region of the Iberian Peninsula. The sample set consisted of 182 samples of leaves of deciduous and evergreen species, showing a wide range of concentrations determined by reference methods: 6.60–35.2 g kg⁻¹ (N), 15.5–66.0% (NDF), 10.2–57.3% (ADF), 3.45–27.4% (lignin) and 5.79–31.3% (cellulose). Reflectance spectra, obtained for samples of dried and ground leaves, were recorded as log1/R (R=reflectance) from 1,100 to 2,500 nm. NIRS calibrations were developed using multiple linear (MLR) and partial least-squares (PLSR) regressions, and tested by external validation. Spectral data were transformed to the first and second derivative (1D, 2D). The PLSR method and derivative transformations provided the best statistics and showed lower standard errors of calibration (SEC) and higher coefficients of multiple determination (R ²). In the external validation the standard errors of prediction (SEP) were 0.76 g kg⁻¹ (N), 2.11% (NDF), 1.47% (ADF), 0.85% (lignin) and 0.86% (cellulose). The results obtained show that NIRS is very effective for the estimation of these organic constituents in leaf tissue of woody species. This technique can be used in ecological or ecophysiological studies as an alternative to the more time-consuming standard methods.

Keywords: Near-infrared reflectance spectroscopy (NIRS); Organic constituents; Leaf tissue; Woody species

Comparative study between a commercial and a homemade capillary electrophoresis instrument for the simultaneous determination of aminated compounds by induced fluorescence detection by Silvia Casado-Terrones; Sonia Cortacero-Ramírez; Alegría Carrasco-Pancorbo; Antonio Segura-Carretero; Alberto Fernández-Gutiérrez (pp. 1835-1847).

The performance of two capillary electrophoresis (CE) instruments, one commercial and one homemade device, were compared for the determination of derivatised aminated compounds with fluorescein isothiocyanate (FITC). The commercial CE system first uses an argon ion laser as excitation source; the homemade CE device uses an inexpensive blue-light-emitting diode (LED) as the light source and a charge-coupled device (CCD) as the detection system. After fine optimisation of several separation parameters in both devices, a co-electroosmotic flow CE methodology was achieved in coated capillary tubing with 0.001% hexadimetrine bromide (HDB), and 50 mmol L⁻¹ sodium borate at pH 9.3 with 20% 2-propanol for the determination of several amines and aminoacids. Analytical performances, applicability in beer samples and other aspects such as cost or potential for miniaturization have been compared for both devices.

Keywords: Capillary electrophoresis; Laser-induced fluorescence; Light-emitting diode (LED)-induced fluorescence; Aminated compounds; Co-electroosmotic flow

Quantitative analysis of enzymatic assays using indoxyl-based substrates by Pablo Fanjul-Bolado; María Begoña González-García; Agustín Costa-García (pp. 1849-1854).

Hydrolysis of indoxyl-based substrates by hydrolytic enzymes is a commonly used semiquantitative detection system that generates a water-insoluble indigo dye which is difficult to quantify. This work describes the quantitative analysis and enzyme kinetics for alkaline phosphatase (AP) and 5-bromo-4-chloro-3-indoxyl phosphate (BCIP) in solution obtained by applying known solubilization methodology from the textiles industry to the enzymatic product. This proposal is based on the reduction of the tetrahalo-indigo blue dye in a basic medium with the aim of generating its aqueous-soluble parent compound termed indigo white, which gives a rich yellow color in solution and is fluorescent. A quantitative ELISA (where a soluble end product is required) is accomplished for first time using BCIP as substrate.

Keywords: Alkaline phosphatase; 5-Bromo-4-chloro-3-indoxyl phosphate; Quantitative assays

Determination of major, minor and trace elements in cobalt-substituted lithium nickelate ceramic powders by inductively coupled plasma optical emission spectrometry by Yodalgis Mosqueda; Mario Pomares; Eduardo L. Pérez-Cappe; Avelina Miranda; Juan C. Fariñas; María T. Larrea (pp. 1855-1862).

An analytical method was developed for the determination of three major (Li, Ni and Co) and fourteen minor or trace elements (Al, Ba, Ca, Cu, Cr, Fe, K, Mg, Mn, Na, Si, Sr, Ti and V) in LiNi1−xCo xO₂ (x = 0.2–0.8) ceramic powders by inductively coupled plasma optical emission spectrometry. Sample dissolution was achieved by 25% nitric acid digestion in a microwave oven. For each element, an analytical line free from spectral interferences was selected. A detailed study of matrix effects over a wide interval of total excitation energy (TEE) lines (1.62–16.50 eV) was performed at near-robust plasma conditions. A remarkable enhancement in atomic lines with TEE <4 eV was noticed, whereas a significant reduction in atomic and ionic lines with TEE >4 eV was observed. The extrapolation to infinite dilution method was successfully used to overcome these nonspectroscopic interferences. Detection limits (3σ) varied from 0.21 mg kg⁻¹ for Sr to 49.7 mg kg⁻¹ for Na. The precision of determination (obtained as the relative standard deviation) was lower than 1% for the major elements Li, Ni and Co and between 0.69 and 10% for minor and trace elements. The accuracy of the method ranged from 91 to 101% for major elements, and from 90 to 110%, or close to this range, for most of the impurities in both of the samples studied.

Keywords: Cobalt-substituted lithium nickelate ceramic; ICP–OES; Elemental determination; Matrix effect correction; Extrapolation to infinite dilution method

Rapid analysis of the essential oils from dried Illicium verum Hook. f. and Zingiber officinale Rosc. by improved solvent-free microwave extraction with three types of microwave-absorption medium by Ziming Wang; Lu Wang; Tiechun Li; Xin Zhou; Lan Ding; Yong Yu; Aimin Yu; Hanqi Zhang (pp. 1863-1868).

A new method of extracting essential oils from dried plant materials has been studied. By adding a microwave-absorption medium (MAM) to a reactor, solvent-free microwave extraction (SFME) was improved and can be used to extract essential oils from dried plant material without pretreatment. With a microwave irradiation power of 85 W it took only approximately 30 min to extract the essential oils completely. The whole extraction process is simple, rapid, and economical. Three types of MAM, iron carbonyl powder (ICP), graphite powder (GP), and activated carbon powder (ACP), and two types of dried plant material, Illicium verum Hook. f. and Zingiber officinale Rosc., were studied. The results were compared with those obtained by use of conventional SFME, microwave-assisted hydrodistillation (MAHD), and conventional hydrodistillation (HD), and the conclusion drawn was that improved SFME was a feasible means of extracting essential oils from dried plant materials, because there were few differences between the composition of the essential oils extracted by improved SFME and by the other methods.

Keywords: Solvent-free microwave extraction; Illicium verum Hook. f.; Zingiber officinale Rosc.; Microwave absorption medium; Essential oil

Development of a multianalyte method for the determination of anabolic hormones in bovine urine by isotope-dilution GC–MS/MS by C. S. Aman; A. Pastor; G. Cighetti; M. de la Guardia (pp. 1869-1879).

A sensitive, specific and selective multianalyte GC–MS/MS method has been developed for the determination of 11 anabolic hormones in bovine urine. After adjusting the urine pH to 4.8, the samples were spiked with deuterated internal standards and submitted to enzymatic hydrolysis with β-glucuronidase/arylsulfatase. Hormones were eluted with methanol through a C18 solid phase cartridge and submitted to a liquid–liquid extraction. Analytes were derivatized by adding N,O-bis(trimethylsilyl)trifluoroacetamide (BSTFA) with 1% trimethylchlorosilane and GC–MS data were obtained in the positive electron impact tandem mass mode. Under these conditions, no matrix effects were observed and limit of detection values were in the range of 0.005 ng/mL (diethylstilbestrol) to 0.38 ng/mL (17α-methyltestosterone and 17α-ethynylestradiol). Recoveries from 81% (α-zeranol) to 149% (17α-methyltestosterone) were found under the selected conditions. These results were better than those found using heptafluorobutyric anhydride (HFBA) as derivative reagent and those measured in full scan and selective ion monitoring modes.

Keywords: Anabolic hormones; Bovine urine; GC–ID–MS/MS; Multiresidual analytical method

Simultaneous determination of five synthetic antioxidants in edible vegetable oil by GC–MS by Lan Guo; Ming-Yong Xie; Ai-Ping Yan; Yi-Qun Wan; Yu-Mei Wu (pp. 1881-1887).

A simple, quick and nontoxic analytical method for the simultaneous determination of five synthetic antioxidants [t-butyl-4-hydroxyanisole (BHA), 2,6-di-t-butyl-hydroxytoluene (BHT), t-butyl hydroquinone (TBHQ), ethoxyquin (EQ) and 2,6-di-tert-butyl-4-hydroxymethyl-phenol (Ionox 100)] in edible vegetable oil has been developed. The analytes were extracted by ethanol, then separated and detected by GC–MS. Extraction conditions such as volume of ethanol required, mixing time and number of extractions were investigated and optimized by an orthogonal array experimental design. The five compounds behaved linearly in the 0.100∼20.0 mg/L concentration range, and the limits of detection (LOD) for BHA, BHT, TBHQ, EQ and Ionox-100 were 1.00, 0.92, 11.5, 0.83 and 1.39 μg/L, respectively. The recoveries at the tested concentrations of 1.00, 20.0 and 100 mg/kg were 75.6∼123%, with coefficients of variation <10.0%. The proposed procedure was successfully applied to the simultaneous analysis of the five antioxidants in soybean oil, tea oil, edible blended oil, rap oil, peanut oil, peanut blended oil and sesame oil samples purchased from local supermarkets.

Keywords: Synthetic antioxidants; Edible vegetable oil; Simultaneous determination; GC-MS

Monitoring lipase-catalyzed butterfat interesterification with rapeseed oil by Fourier transform near-infrared spectroscopy by Hong Zhang; Huiling Mu; Xuebing Xu (pp. 1889-1897).

This work demonstrates the application of FT-NIR spectroscopy in order to monitor the enzymatic interesterification process for butterfat modification. The reactions were catalyzed by Lipozyme TL IM at 70 °C for the blend of butterfat/rapeseed oil (70/30, w/w) in a packed-bed reactor. The blend and interesterified fat samples were measured in liquid form at 70 °C by transmission mode-based FT-NIR over the spectral region 12000–4000 cm⁻¹. The calibration of FT-NIR for conversion degree (evaluated by the triglyceride profile, which was represented by the triglyceride peak ratio) and solid fat content (SFC) of the interesterified products was carried out using partial least squares (PLS) regression. Good correlations were observed between the NIR spectra and ln (peak ratio), and between the NIR spectra and the SFC at 5 °C over the spectral range 5269–4513 cm⁻¹. Overall, transmission-mode FT-NIR spectroscopy performed at 70 °C yielded conditions close to those used during the interesterification process, implying that this method could be used to control the enzymatic interesterification process online.

Keywords: FT-NIR; Enzymatic interesterification; Monitoring; Conversion degree; Packed-bed reactor

Electrochemical determination of sugars by use of multilayer thin films of ferrocene-appended glycogen and concanavalin A by Katsuhiko Sato; Daisuke Kodama; Jun-ichi Anzai (pp. 1899-1904).

Multilayer thin films containing concanavalin A (Con A) and ferrocene-appended glycogen (FcGly) were prepared by a layer-by-layer deposition Con A and FcGly by biological affinity (lectin–sugar interaction) on a glassy-carbon electrode. The electrochemical response of the Con A–FcGly film-coated electrode to sugars was investigated. A cyclic voltammogram (CV), typical of redox species confined to the surface of the electrode, was obtained. The peak current (resulting from the electric charge involved in the redox reaction) in the CV from the electrode decreased on addition of sugars in the solution, because the amount of FcGly on the electrode surface decreased as a result of disintegration of the Con A–FcGly film on addition of sugar. Thus, d-glucose and other sugars at millimole per liter levels can be detected by use of Con A–FcGly films-coated electrodes.

Keywords: Concanavalin A; Glycogen; d-Glucose detection; Multilayer thin film

The electrochemical behavior of p-benzenediol on a self-assembled monolayers Pt electrode modified with N-(2-mercapto-1,3,4-thiadiazol-5-yl)-N′-(4-substituted-arylacetyl) urea by Chuan-yin Liu; Ji-ping Yao; Hong-wu Tang; Sheng-ping Zhu; Jun-fu Hu (pp. 1905-1911).

Two novel N-(2-mercapto-1,3,4-thiadiazol-5-yl)-N′-(4-substituted-arylacetyl) urea compounds have been synthesized, characterized by NMR and MS, and used as self-assembly reagents to form self-assembled monolayers (SAMs) on Pt electrodes. The modified electrodes were characterized by electrochemical methods. The electrochemical behavior of p-benzenediol at the SAMs electrodes was investigated. It was found that the electrochemical response to p-benzenediol is controlled by diffusion and can be electrocatalyzed to obtain more symmetrical redox peaks and higher voltammetric current response at the SAMs electrodes, with a peak separation of 80 mV. For p-benzenediol the process at the SAMs electrodes is quasi-reversible with a rate constant of 0.6742 s⁻¹. The SAMs electrodes have been used to determine p-benzenediol by differential pulse voltammetry. The peak current was linear for concentrations of p-benzenediol in the range 1×10⁻⁷−5×10⁻⁴ mol L⁻¹ and the detection limit was 4.0×10⁻⁸ mol L⁻¹. The SAMs electrodes were used to determine p-benzenediol in real photographic developer and in a synthetic waste water sample; the standard addition recovery was in the range 96.6–100.4%.

Keywords: Self-assembled monolayers; p-Benzenediol; Electrocatalysis; Quasi-reversible process

Electrochemical studies of (−)-epigallocatechin gallate and its interaction with DNA by Xiaofeng Zheng; Anyue Chen; Tomonori Hoshi; Jun-ichi Anzai; Genxi Li (pp. 1913-1919).

In this paper, an electrochemical investigation of (−)-epigallocatechin gallate (EGCG) and its interaction with DNA is presented. Via an electrochemical approach assisted by ultraviolet–visible (UV–Vis) spectroscopy, we propose that EGCG can intercalate into DNA strands forming a nonelectroactive complex, which results in the decrease of the anodic peak current of EGCG. Meanwhile, an electrochemical study with the DNA–Cu(II)–EGCG system shows that damage to DNA can be recognized electrochemically via the increase in the anodic peak current resulting from the oxidation of guanine and adenine bases. The damage can also be recognized spectrophotometrically via an increase in the 260 nm absorption band. In addition, it was found that EGCG is able to discriminate dsDNA from ssDNA, making a potential electrochemical indicator for the detection of DNA hybridization events. A rapid and convenient method of detecting EGCG was also developed in this work. Figure Interaction of EGCG with DNA and damage to DNA in the presence of Cu(II)

Keywords: (−)-Epigallocatechin gallate; DNA; Interaction

A new approach to flow-batch titration. A monosegmented flow titrator with coulometric reagent generation and potentiometric or biamperometric detection by Emerson Vidal de Aquino; Jarbas José Rodrigues Rohwedder; Celio Pasquini (pp. 1921-1930).

Monosegmented flow analysis (MSFA) has been used as a flow-batch system to produce a simple, robust, and mechanized titrator that enables true titrations to be performed without the use of standards. This paper also introduces the use of coulometry with monosegmented titration by proposing a versatile flow cell. Coulometric generation of the titrand is attractive for titrations performed in monosegmented systems, because the reagent can be added without increasing the volume of sample injected. Also, biamperomeric and potentiometric detection of titration end-points can increase the versatility of the monosegmented titrator. The cell integrates coulometric generation of the titrand with detection of end-point by potentiometry or biamperometry. The resulting titrator is a flow-batch system in which the liquid monosegment, constrained by the interfaces of the gaseous carrier stream, plays the role of a sample of known volume to be titrated. The system has been used for determination of ascorbic acid, by coulometric generation of I₂ with biamperometric detection, and for determination of Fe(II), by coulometric generation of Ce(IV) with potentiometric detection of the end-point, both in feed supplements.

Keywords: Flow Titration; Flow-Batch; Coulometry; Potentiometry; Iron(II); Ascorbic Acid; Monosegmented system

Analysis of carbamazepine and its active metabolite, carbamazepine-10,11-epoxide, in human plasma using high-performance liquid chromatography by Eun kyung Oh; Eunmi Ban; Jong Soo Woo; Chong-Kook Kim (pp. 1931-1936).

A sensitive method based on high-performance liquid chromatography (HPLC) with ultraviolet (UV) detection was developed for the determination of carbamazepine (CBZ) and one of its active metabolites, carbamazepine-10,11-epoxide (CBZ-E) in human plasma. CBZ, CBZ-E and the internal standard (IS) 10,11-dihydrocarbamazepine were extracted from human plasma into methyl tert-butyl ether. CBZ, CBZ-E and the IS were successfully separated on an RP C18 column with a mobile phase of acetonitrile:methanol:water (18:19:63, v/v/v) and monitored via UV detection at 210 nm. The calibration curves were linear over the concentration ranges of 0.01–10 μg/mL for CBZ and 0.005–5 μg/mL for CBZ-E in human plasma, respectively. The method displayed excellent sensitivity, precision and accuracy, and was successfully applied to the quantification of CBZ and CBZ-E in human plasma after oral administration of a single 200 mg CBZ CR tablet. This method is suitable for bioequivalence studies following single doses given to healthy volunteers.

Keywords: Carbamazepine; Carbamazepine-10,11-epoxide; High-performance liquid chromatography (HPLC); Bioequivalence study

Preparative HPLC separation of bambuterol enantiomers and stereoselective inhibition of human cholinesterases by Ivana Gazić; Anita Bosak; Goran Šinko; Vladimir Vinković; Zrinka Kovarik (pp. 1937-1937).

Sections

Personal tools

Analytical and Bioanalytical Chemistry (v.386, #6)

Sections

Personal tools

Local resources

Analytical and Bioanalytical Chemistry (v.386, #6)