Preparation and characterization of molecularly imprinted electropolymerized carbon electrodes

Molecularly imprinted polymers (MIPs) selective for fluorescein, rhodamine or 2,4-dichlorophenoxyacetic acid (2,4-D) were electropolymerized onto graphite electrodes using an aqueous solution equimolar in resorsinol/ortho-phenylenediamine and in the presence of the template molecule. For the dyes, the MIP-coated electrodes showed higher affinity for their template molecule than for a non-template dye. The 2,4-D-MIP-coated electrode showed a concentration dependent response for 2,4-D as compared to the polymer-coated electrode prepared in the absence of template molecule.     

Fabrication and performance of a three-dimensionally adjustable device for the amperometric detection of microchip capillary electrophoresis

A microchip CE-amperometric detection (AD) system has been fabricated by integrating a two-dimensionally adjustable CE microchip and an AD cell containing a one-dimensionally adjustable disk detection electrode in a Plexiglas holder. It facilitates the precise 3-D alignment between the channel outlet and the detection electrode without a complicated 3-D manipulator. The performance of this unique system was demonstrated by separating five aromatic amines (1,4-phenyldiamine, aniline, 2-methylaniline, 4-chloroaniline, and 1-naphthylamine) of environmental concern. Factors influencing their separation and detection processes were examined and optimized. The five analytes have been well separated within 140 s in a 74 cm long separation channel at a separation voltage of +2500 V using a 10 mM phosphate buffer (pH 3.5). Highly linear response is obtained for the five analytes over the range 20-200 microM with the detection limits ranging from 0.46 to 1.44 microM, respectively. The present system demonstrated long-term stability and reproducibility with RSDs of less than 5% for the peak current (n = 9). The new approach for the microchannel-electrode alignment should find a wide range of applications in CE, flowing injection analysis, and other microfluidic analysis systems.     

Determination and Differentiation of Two Seemingly Identical Medicinal Herbs by Capillary Electrophoresis with Electrochemical Detection

A high-performance capillary electrophoresis with electrochemical detection (CE-ED) method has been employed for the determination of six bioactive ingredients in traditional Chinese herbs, Herba cepbalanoplosis segeti and Herba cirsii japonici. The effects of several factors such as the acidity and concentration of running buffer, the separation voltage, the applied potential and the injection time on CE-ED were investigated. Under the optimum conditions, the six analytes could be well separated within 21 min in a 75 cm length capillary at the separation voltage of 15 kV in a 50 mmol L^–1 borax running buffer (pH 8.4). A 300 m diameter carbon disk electrode was used as the working electrode positioned carefully opposite the outlet of the capillary in a wall-jet configuration at potential of +950 mV (vs. SCE). Good linear relationship was established between peak current and concentration of analytes over two orders of magnitude with detection limits (S/N=3) ranged from 1.5×10^–7 to 6.0×10^–7 g mL^–1 for all six analytes. This proposed method has been successfully applied for the analysis of traditional Chinese herbs after a relatively simple extraction procedure, further on, for the differentiation of these above two seemingly identical herbs based on their electropherograms or characteristic electrochemical profiles.     

Fluorescence-based nucleic acid detection and microarrays

Recent analytical innovations for nucleic acid detection have revolutionized the biological sciences. Single nucleic acid sequence detection methods have been expanded to incorporate multiplexed detection strategies. A variety of nucleic acid detection formats are now available that can address high throughput genomic interrogation. Many of these parallel detection platforms or arrays, employ fluorescence as the signaling method. Fluorescence-based assays offer many advantages, including increased sensitivity, safety and multiplexing capabilities, as well as the ability to measure multiple fluorescence properties. Multiplexed microarray platforms provide parallel detection capabilities capable of measuring thousands of simultaneous responses. This review will discuss both single target detection and microarray applications with a focus on gene expression and pathogenic microorganism (PM) detection.     

Selectivity enhancement of a metal-sensitive flame ionization detector for capillary gas chromatography

A novel modification of a hydrogen-atmosphere flamo ionization detector (HAFID) is presented which attenuates response to hydrocarbon compounds, significantly enhancing selectivity towards organometallic compounds by more than an order of magnitude. Chromatograms of an organometallic compound test mixture and regular leaded gasoline are presented to depict the specificity of the response.     

Validation of HPLC Analysis of 2-Phenoxyethanol, 1-Phenoxypropan-2-ol, Methyl, Ethyl, Propyl, Butyl and Benzyl 4-Hydroxybenzoate (Parabens) in Cosmetic Products, with Emphasis on Decision Limit and Detection Capability

The Commission Decision of August 12, 2002 on the performance of analytical methods and the interpretation of results was applied to the HPLC method for the analysis of parabens, 2-phenoxyethanol and 1-phenoxypropan-2-ol in cosmetic products. This method is published in the seventh Directive 96/45/EC of the European Commission. Non-compliant concentrations, taking into account the data distribution (CC) and the probability of false negative values (CC) were determined. The repeatability and reproducibility amount to <4% and <7%, respectively. These values were obtained with blanc samples that were fortified in the laboratory. Calibration linearity was confirmed by absence of lack of fit for all seven preservatives. Matrix effects on the determinations of the preservatives in body milk or shampoo are negligible.     

Some observations on the automation by flow injection analysis of the spectrophotometric determination of amino acids and proteins witho-phthalaldehyde

Automation by flow injection analysis with Spectrophotometric detection of the determination of total amino acids and proteins witho-phthalaldehyde is not straightforward. The use of spectrophotometry, instead of spectrofluorimetry, and of N-acetyl-L-cysteine, instead of the conventional mercaptoethanol is advantageous because of the lower variability of absorptivities with respect to fluorescence yields, and the larger stability of the derivatives. Under adequate working conditions and with leucine as reference, the procedure can be used for the evaluation of total amino acids. A similar procedure is proposed for the analysis of proteins in a sample. Limits of detection are 1 × 10^–5 M for amino acids, and 1 × 10^–6 M for proteins, respectively.     

Improved method for the determination of kynurenic acid in rat plasma by column-switching HPLC with post-column fluorescence detection

Kynurenic acid (KYNA), an endogenous antagonist of ionotropic glutamate and α7 nicotinic receptors, was fluorometrically determined by column-switching high-performance liquid chromatography (HPLC) with fluorescence detection. The HPLC system consists of two octadecyl silica (ODS) columns, both of which are connected with an anion-exchange column (trapping column). Following sample injection onto the HPLC column, KYNA was separated on the first ODS column with a mobile phase of H₂O/acetonitrile (95/5) containing 0.1% acetic acid. The peak fraction of KYNA was trapped on the anion-exchange column by changing the position of a six-port valve and then introduced into the second ODS column. Subsequently, KYNA was detected fluorometrically as a fluorescence complex formed with zinc ion which was pumped constantly. Instrumental limit of detection was approximately 0.16 nM, which corresponded to 8.0 fmol (per 50 μl injection, signal to noise ratio 3), and the limit of quantification was 0.53 nM (signal to noise ratio 10). Intra- and inter-day relative standard deviations were 1.1-3.9% (n = 3) and 3.0-5.3% (n = 3), respectively. The peak of KYNA in rat plasma was clearly detected by the proposed column-switching HPLC system after a facile pretreatment procedure. Intra- and inter-day relative mean errors were −1.6-1.4% (n = 3) and −2.4 to −0.4% (n = 3), respectively, with a satisfactory precision (within 5.0%). A calibration curve for the determination of KYNA showed a good linearity (r² > 0.999) in the range of 25-200 nM. The KYNA concentrations in the plasma of male Sprague-Dawley rats (8-week-old) were 44 ± 5.5 nM (mean ± S.E., n = 5). In ketamine-treated rats, which are animal models of schizophrenia, the plasma KYNA concentrations were significantly increased compared with those in the control rats (p < 0.05).     

EPR detection of foods preserved with ionizing radiation

The applicability of the epr technique for the detection of dried vegetables, mushrooms, some spices, flavour additives and some condiments preserved with ionizing radiation is discussed. The epr signals recorded after exposure to gamma rays and to beams of 10 MeV electrons from linac are stable, intense and specific enough as compared with those observed with nonirradiated samples and could be used for the detection of irradiation. However, stability of radiation induced epr signals produced in these foods depends on storage condition.

No differences in shapes (spectral parameters) and intensities of the epr spectra recorded with samples exposed to the same doses of gamma rays (⁶⁰Co) and 10 MeV electrons were observed     

Fluorescence immunological determination of immunoglobulin G in human serum by high-performance liquid gel-permeation chromatography

Summary A high-performance liquid gel-permeation chromatographic method is described for the determination of human serum immunoglobulin G (IgG) by separating the fluorescent immuno complex from the free fluorescence-labeled antibody. Fluorescence-labeled antibody used in this study was fluorescein isothiocyanate (FITC)-labeled Fab fragment goat anti-human IgG (anti-IgG Fab). Immuno complexes and antibody of different molecular sizes can be separated. FITC-labeled anti-IgG Fab was added to the serum and the mixture is passed through the column. An immuno complex separates as well-delineated peak in the column void volume, and was measured by the fluorescence of the column eluate (Ex=490nm, Em=520nm). The total analysis time for a serum sample was approximately 15min. The minimum detection limit was 25 mg/dl. The relative standard deviation was below 2% (peak area). The results of the HPL-GPC analysis correlate well with those obtained by laser nephelometric assay (r=0.992).