Analytica Chimica Acta (v.521, #2)

A dual fluorophore system for simultaneous bioassays by Christel Hempen; Uwe Karst (117-122).
A detection scheme for the simultaneous evaluation of two bioassays based on fluorescence spectroscopy is presented. For the determination of hydrogen peroxide-generating enzymes or peroxidases, the non-fluorescent 4-(N-methylhydrazino)-7-nitro-2,1,3-benzooxadiazole (MNBDH) is converted to the strongly fluorescent 4-(N-methylamino)-7-nitro-2,1,3-benzooxadiazole (MNBDA). Phosphatases are detected based on the cleavage of the non-fluorescent 5-fluorosalicyl phosphate (5-FSAP) under formation of the fluorescent 5-fluorosalicylic acid (5-FSA). While excitation of the fluorophores may be carried out at the same wavelength, their emission spectra differ significantly. This allows the read-out of both assays using commercially available microplate readers without additional chemometric tools. Compared with individual assays, limits of detection are similar, and linearity of the calibration functions for both enzymes is observed over 2–3 concentration decades starting at the limit of quantification. The simultaneous determination of glucose oxidase and acid phosphatase in honey is presented as example for the application of the detection scheme.
Keywords: Bioassays; Fluorescence; Simultaneous determinations; Glucose oxidase; Peroxidase; Phosphatase;

Multi-elemental characterization of soft biological tissues by inductively coupled plasma–sector field mass spectrometry by Emma Engström; Anna Stenberg; Svetlana Senioukh; Roland Edelbro; Douglas C Baxter; Ilia Rodushkin (123-135).
The performance of double-focusing, sector field ICP–MS (ICP–SFMS) for the multi-elemental analysis of soft tissues following microwave-assisted digestion with nitric acid was evaluated and factors affecting method limits of detection discussed. Accuracy was assessed by replicate analyses of certified reference materials and by participation in performance evaluation programs; the precision was better than 5% relative standard deviation (RSD) for the majority of elements. Cl was the only element for which ICP–SFMS data consistently deviated from certified concentrations in the reference materials tested. Comparison between results obtained by ICP–SFMS and ICP optical emission spectrometry showed good agreement for elements present in tissues at concentrations above 2 μg g−1. The concentrations of 68 elements in different fish and animal soft tissues (muscle, liver, kidney, lung and brain) are presented, and, where possible, compared with previously published data.
Keywords: Biological tissues; Inductively coupled plasma–sector field mass spectrometry; Multi-elemental analysis; Digestion;

Determination of platinum and rhodium in dust and plant samples using microwave-assisted sample digestion and ICP-MS by Matti Niemelä; Paavo Perämäki; Juha Piispanen; Jarmo Poikolainen (137-142).
The platinum group elements (PGEs), particularly platinum, palladium and rhodium, are nowadays increasingly emitted into the environment from automotive catalytic converters. Thus, a method for the determination of PGEs (especially platinum and rhodium) in dust and plant samples was developed. The developed method was based on microwave-assisted sample digestion and inductively coupled plasma mass spectrometric (ICP-MS) determination. Spectral interferences in ICP-MS determination were corrected using mathematical correction equations based on signal ratio measurement. In addition, platinum and rhodium concentrations in the digested dust samples were also determined after Te coprecipitation without correction of the interferences. The results for platinum and rhodium in reference materials (NIST SRM 2557, recycled monolith autocatalyst and BCR-723, road dust) were in good agreement with the certified values. Preliminary results for the anthropogenic platinum and rhodium emissions in Oulu, northern Finland, based on dust and plant samples, indicated a common traffic-related source of these metals.
Keywords: Platinum group elements; ICP-MS; Sample preparation; Microwave digestion; Dust samples; Plant samples; Moss;

Near-infrared spectroscopy was used to control an esterification reaction between glycerine and middle- or long-chain fatty acids performed in a laboratory-scale reactor. The process involves the initial formation of monoglycerides, which is followed by that of di- and triglycerides as well as transesterification. Establishing the end point of the process is critical with a view to ensuring that the end product will have the composition required for its intended use. PLS calibration was applied to industrial and laboratory-scale batch samples, and laboratory samples were additionally used to extend calibration ranges and avoid correlation between the concentration of the batch samples. In this way, PLS calibration models for glycerine, fatty acids, water, and mono-, di- and triglycerides, were constructed. The proposed method allows the reaction to be monitored in real time, thereby avoiding long analysis times, excessive reagent consumption and the obtainment of out-of-specification products.
Keywords: NIR; Control esterification process; Multivariate calibration;

Sweeteners determination in table top formulations using FT-Raman spectrometry and chemometric analysis by Sergio Armenta; Salvador Garrigues; Miguel de la Guardia (149-155).
A partial least squares (PLS) Fourier transform Raman spectrometry procedure based on the measurement of solid samples contained inside standard glass vials, has been developed for direct and reagent-free determination of sodium saccharin and sodium cyclamate in table top sweeteners. A classical 22 design for standards was used for calibration, but this system provides accuracy errors higher than 13% w/w for the analysis of samples containing glucose monohydrate. So, an extended model incorporating glucose monohydrate (23 standards) was assayed for the determination of sodium saccharin and sodium cyclamate in all the samples. Mean centering spectra data pre-treatment has been employed to eliminate common spectral information and root mean square error of calibration (RMSEC) of 0.0064 and 0.0596 was obtained for sodium saccharin and sodium cyclamate, respectively. A mean accuracy error of the order of 1.1 and 1.9% w/w was achieved for sodium saccharin and sodium cyclamate, in the validation of the method using actual table top samples, being lower than those obtained using an external monoparametric calibration. FT-Raman provides a fast alternative to the chromatographic method for the determination of the sweeteners with a three times higher sampling throughput than that obtained in HPLC. On the other hand, FT-Raman offers an environmentally friendly methodology which eliminates the use of solvents. Furthermore, the stability of samples and standards into chromatographic standard glass vials allows their storage for future analysis thus avoiding completely the waste generation.
Keywords: Sweeteners; Sodium saccharin; Sodium cyclamate; Raman; Powder analysis;

Chelating resin from functionalization of chitosan with complexing agent 8-hydroxyquinoline: application for metal ions on line preconcentration system by Amarildo Otavio Martins; Edson Luiz da Silva; Eduardo Carasek; Norberto S Gonçalves; Mauro C.M Laranjeira; Valfredo T de Fávere (157-162).
This study describes the functionalization of biopolymer chitosan, using the complexing agent 8-hydroxyquinoline (oxine) by reaction of diazotization. The chelating resin was characterized by degree of deacetylation, infrared, Raman spectroscopy. The efficiency of the chelating resin and accuracy of the proposed method was evaluated by the metal ion recovery technique in the analysis of potable water, lake water, seawater and a certified sample of oyster tissue. The metal ions Cd(II) and Cu(II) in the samples were previously enriched in a minicolumn and flow injection flame atomic absorption spectrometry (FI-FAAS) determined the concentrations of the analytes. The chelating resin exhibited high selectivity for Cd(II) at pH 7 and for Cu(II) at pH 10. The eluent concentration was tested by the use of HNO3 in concentrations of 0.1–3 mol l−1 maximum response was obtained at 0.5 mol l−1 for Cd(II) and Cu(II), with R.S.D. values of 0.4%. The analytes gave relative standard deviations (R.S.D.) of 1.5 and 0.7% for solutions of Cd(II) and Cu(II), respectively (n = 7) containing 20 μg l−1 of the metal ions, defining a high reproducibility. The limits of detection (LOD) were 0.1 μg l−1 for Cd(II) and 0.4 μg l−1 for Cu(II). The analytical properties of merit were obtained using the parameters previously optimized with preconcentration time of 90 s. The chelating resin showed chemical stability within a wide range of pH and the efficiency was not altered for the preconcentration of the metal ions during all the experiments.
Keywords: Chelating resin; Chitosan; 8-Hydroxyquinoline; Preconcentration;

Calix[6]arene acetic acid extraction behavior and specificity with respect to nucleobases by Kojiro Shimojo; Tatsuya Oshima; Masahiro Goto (163-171).
The extraction behavior of various nucleobases was investigated with a series of calixarene derivatives. A calix[6]arene acetic acid was found to extract around 80% of adenine from an aqueous phase, and showed the highest specificity for adenine among all the nucleobases tested. The important factors influencing the recognition properties of calixarene were discovered to be its cyclic structure, its cavity size, and the functional carboxylic acid groups. The carbonyl group and the position of the amino group in the guest molecules also affect the extraction efficiency. Calix[6]arene forms a stable 1:1 complex with adenine by entrapping it into the cavity. Adenine was readily recovered from the organic phase by contacting with a fresh acidic solution.
Keywords: Solvent extraction; Calixarene; Nucleobase; Inclusion phenomena; Molecular recognition;

In this work a new cloud point extraction (CPE) methodology was developed for the separation and preconcentration of cadmium and nickel. The analyte in the initial aqueous solution was complexed with dithizone and Triton X-114 was added as surfactant. After phase separation, based on the cloud point of the mixture, and dilution of the surfactant-rich phase with tetrahydrofuran (THF), the enriched analytes were determined by flame atomic absorption spectrometry. After optimization of the complexation and extraction conditions and preconcentration of only 10 ml of sample in the presence of 0.05% Triton X-114, the enhancement factors of 52 and 39 and the detection limits of 0.31 μg l−1 and 1.2 μg l−1 were obtained for cadmium and nickel respectively. The proposed method was applied satisfactorily to the determination of cadmium and nickel in water samples.
Keywords: Cloud point; Triton X114; Dithizone; Cd and Ni determination;

Extraction and quantification of antioxidants from low-density polyethylene by microwave energy and liquid chromatography by M.S Dopico Garcia; J.M López V; R Bouza; M.J Abad; E González Soto; M.V González Rodrı́guez (179-188).
In this paper a experimental design is applied to optimize the quantification of hindered phenol Irganox 1076, phosphite antioxidant Irgafos 168 and their oxidized product tri[2,4-di-tert-butylphenyl]phosphate from low-density polyethylene (LDPE). The developed analytical method consists of two steps: microwave-assisted extraction and reversed-phase liquid chromatography (LC) coupled with ultraviolet diode-array detector. A Plackett–Burman design was carried out in order to find the significant experimental parameters affecting the antioxidants extraction by microwave energy. These parameters were subsequently optimized by a central composite design. The performed method allows extracting the studied antioxidants at low temperature in a short time without degradation of phosphite antioxidant Irgafos 168.
Keywords: Antioxidants; Experimental design; Microwave; LC; LDPE;

Treatment of turkeys with nitroimidazoles by J Polzer; C Stachel; P Gowik (189-200).
In several animal studies turkeys were treated with different nitroimidazoles (Dimetridazole, Metronidazole, Ronidazole, Ipronidazole). After slaughtering, different matrices (breast muscle, leg muscle, liver, plasma, retina) were analysed for their analyte content, for the percentage of hydroxy-metabolites, for homogeneity, stability and bound, and conjugated residues. The tests showed that for animals treated with Dimetridazole and Ipronidazole, the hydroxy-metabolites (2-hydroxymethyl-1-methyl-5-nitroimidazole (HMMNI) and 1-methyl-2-(2′-hydroxyisopropyl)-5-nitroimidazole (IPZOH)) are the relevant target analytes, whereas for animals treated with Ronidazole and Metronidazole, the parent drug itself is the most relevant analyte. In muscle samples an inhomogeneous analyte distribution was found. Degradation studies showed a rapid decline of the analyte concentration in muscle and liver samples stored at room temperature and a decelerated degradation at 4 °C. In plasma and retina samples, however, the analytes were stable during storage under the same conditions. In these matrices the analytes were found to be present in considerably higher concentrations than in muscle or liver and could be detected for a longer period of time after withdrawal of the medication. Therefore, plasma or retina can be recommended as target matrices for the residue control of nitroimidazoles in turkeys.
Keywords: Residue control; Nitroimidazoles; Homogeneity; Stability; Turkey; Retina; Plasma;

The electrochemical behavior of 1-hydroxypyrene at a thin-film mercury electrode was investigated to provide the basis for development of an inexpensive, sensitive, and reliable method for determination of polycyclic aromatic hydrocarbon (PAH) metabolites in biological matrices. Optimum experimental conditions for analytical applications were obtained in 0.01 M NaOH solution using an accumulation potential of −0.20 V, a scan rate of 200 mV s−1, a pulse height of 50 mV, and the square wave scan mode. The electrode response to 1-hydroxypyrene is linear up to 100 ppb. For an accumulation time of 10 min, the detection limit was 0.23 ppb (1.06 × 10−9  mol L−1). The method was applied to the determination of 1-hydroxypyrene in samples of human urine. Cyclic voltammetry and square wave voltammetry were used to characterize the interfacial and redox behavior.
Keywords: 1-Hydroxypyrene determination; Copper; 9-Phenanthrol; Human urine; Polycyclic aromatic hydrocarbons; Thin-film mercury electrode; Square wave stripping voltammetry;

To detect a low level of acetylcholine (Ach) output in the dialysate of rat frontal cortex, osmium-peroxidase redox polymer modified gold-ring disk electrode (Os-HRP-RDE) was fabricated in high-performance liquid chromatography (HPLC)-electrochemical detection (ECD). In comparison with platinum electrode, the sensitivity of acetylcholine detection by Os-HRP-RDE was stable during a day, the coefficient of variation was 0.95%, and the decrease of the sensitivity for 1 week was 5.4%. The Os-HRP-RDE could detect lower than 3 fmol of acetylcholine. The present technique, in the absence of acetylcholinesterase (AchE) inhibitor, is useful to facilitate physiological investigations of cholinergic neuronal activity in the brain.
Keywords: Acetylcholine; HPLC; Os-HRP; Au-ring disk; Microdialysis;

Optimisation procedure for the inhibitive determination of chromium(III) using an amperometric tyrosinase biosensor by O. Domı́nguez Renedo; M.A. Alonso Lomillo; M.J. Arcos Martinez (215-221).
An enzymatic amperometric procedure for measurement of chromium(III), based on the inhibitive action of this metal on tyrosinase enzyme activity, was developed for this purpose. A glassy carbon electrode was used as a support for the electropolymerisation of a polypyrrole (PPy) film, in which tyrosinase was immobilised. The parameters were optimised using experimental design methodology. To determine the optimum conditions and the influence of the factors in the experiment, a canonical analysis of the response surface was carried out. Under these conditions, the repeatability (R.S.D. < 5.8%), the reproducibility (R.S.D. < 8.3%), and the stability of the modified electrode were all analysed. The detection limit for Cr(III) was calculated at a value of 5.0 × 10−7  mol dm−3 with a relative standard deviation (R.S.D.) of 5.8%. Analysis of the possible effect of the presence of foreign ions in the solution was performed. The method was applied to determine levels of chromium(III) in spiked urine samples, in the waste water from a tannery, and in river water from an industrial area.
Keywords: Tyrosinase; Biosensor; Chromium; Experimental design;

Determination of fenoterol hydrobromide by sequential injection analysis with spectrophotometric detection by Negussie W Beyene; Jacobus F van Staden; Raluca I Stefan (223-229).
A rapid, economical and automated sequential injection spectrophotometric determination of the phenolic sympathomimetic drug, fenoterol hydrobromide, is reported. The method is based on the reaction of fenoterol hydrobromide with 4-aminoantipyrine and potassium hexacyanoferrate and the absorbance of the colored product monitored at 505 nm. Chemical as well as physical SIA parameters that affect the signal response have been optimized in order to get better sensitivity, higher sampling rate and better reagent economy. Using the optimized parameters, a linear relationship between the relative peak height and concentration was obtained in the range 0.5–40 mg L−1. The detection limit (as 3σ value) was 0.1 mg L−1 and precision was 1.8 and 1.6% at 2 and 5 mg L−1, respectively. As compared to previous works, wide linear range, low detection limit, and highly economical reagent consumption are the advantages of our method.
Keywords: Flow techniques; Sympathomimetic drugs; Anti-asthmatics; Bronchodilator;

Application of hand scanner in multivariate quantification of povidone-iodine (PVI), as a popular antiseptic agent, in some of pharmaceutical products is presented. Brightness, contrast, and mixed gamma were the adjustable scanner parameters. For selection of optimum values of the scanner parameters, partial least squares (PLS) and multiple linear regression (MLR), coupled with genetic algorithm, were performed. For the selected variables, both MLR and PLS performances were similar and appropriate. From the results obtained, it was concluded that the simpler method of MLR could be successfully applied instead of PLS, which requires more statistical experience. The considered concentration range for PVI in the calibration and prediction samples was 0.0–10.0% (w/v). For the analysis of pharmaceutical samples, generalized standard addition method (GSAM) was applied (on the variables selected by GA) and desirable results were obtained. Relative standard error (RSE) of less than 8% was obtained for the majority of samples analyzed.
Keywords: Povidone-iodine; Partial least squares; Generalized standard addition method;

Spectroscopic information retained in screen photo-assisted techniques by Daniel Filippini; Ingemar Lundström (237-244).
The computer screen photo-assisted technique (CSPT), a recently reported method for characterization of colorimetric assays, has been modeled and the ability of the technique to retain key spectral features has been determined.The robustness of CSPT to operate with different kind of screens is demonstrated and a physical explanation of the result is provided.Simulated CSPT transmittances corroborate experimental results, and enable comparison with standard methods for qualitative and quantitative evaluation.
Keywords: Colorimetric assays; Photo-assisted sensing; (Bio)chemical images; Visible spectroscopy;

Author Index (245-246).