Analytical Letters

Quinn, John G.; O'Kennedy, Richard

doi: 10.1080/00032719908542911pmid: N/A

The field of biomolecular interaction analysis has been revolutionised over the past decade with the development of biosensors that enable 'real-time' monitoring. Characteristically, these biosensing technologies combine mass sensitive transducers, biointerfacial chemistry and the specificity of a wide range of affinity ligands. Considerable development of these elements has occurred over the past few years and promises to create a new generation of portable biosensors. The majority of transducers for 'real-time' biomolecular interaction analysis are generic evanescent wave-based mass sensors or direct mass sensors. The construction of a suitable biointerface is a key component in biosensor design and has a critical influence on the biosensor's performance. Recent developments in these areas are presented with particular emphasis on commercially available technology.

Skládal, Petr; Horácek, Jirí

doi: 10.1080/00032719908542912pmid: N/A

The developed piezoelectric quartz crystal microbalance sensor was compared with the commercial resonant mirror-based optical biosensor IAsys. The former one was based on 10 MHz crystals and flow-through arrangement, the latter employed a stirred cuvette. The same biosensing layers were produced by immobilization of 4,4′-dichlorobiphenyl on aminosilanized surfaces through an albumin bridge. The affinity binding of three corresponding polyclonal antibodies was studied and the kinetic rate constants ka and kd were determined. The same values of constants were obtained for sheep antibody, smaller values of ka (approx. twice) were obtained for the other two lower-affinity antibodies using the piezosensor. In the additional comparison during a competitive assay of atrazine, both transducers were able to distinguish 0 and 0.1 μg/L atrazine, however, the IAsys signal was less noisy. For bioaffinity kinetic studies, the piezoelectric biosensor was found to be an economic alternative to the advanced optical biosensors.

Mori, Hisakazu; Kogure, Mamie; Kuroda, Yohko; Nissato, Miyuki; Tamura, Yukiko; Watanabe, Nahoko; Yamamoto, Haruhiko

doi: 10.1080/00032719908542913pmid: N/A

Simultaneous determination of glucose and sucrose was achieved with an apparatus containing two enzyme reactors and an octadecylsilica column positioned between the two reactors in tandem in a single flow line. The enzymes used were mutarotase and glucose dehydrogenase for glucose analysis, and sucrose phosphorylase, phosphoglucomutase and glucose-6-phosphate dehydrogenase for sucrose analysis. The pH and the concentration of the phosphate buffer used as the medium for immobilization of mutarotase and glucose dehydrogenase to activated aminopropyl glass were studied. Several aqueous buffers and their pH values were also examined for use as the carrier medium, and HEPES buffer (pH 7.5) was the most favorable. In this apparatus under these conditions, NADH due to glucose and that due to sucrose were observed separately. The calibration curve for glucose was linear in the range of 1 - 500 μM, and that for sucrose, 5 - 500 μM. This method was applied to the analyses of glucose and sucrose in several beverages. The results were in good agreement with those obtained by a commercially available test-kit method.

Suye, Shin-ichiro; Miura, Jun'ichiro; Ohue, Masatoshi

doi: 10.1080/00032719908542914pmid: N/A

A sensitive fluorometric method was developed for the determination of urea. The method was based on the hydrolyzation of urea catalyzed by urease with pH and the change of fluorescence intensity of sodium fluorescein as a pH sensitive dye in the enzymatic reaction. The increase in fluorescence intensity of sodium fluorescein was measured at λex= 490 nm and λem= 514 nm. Under the optimum conditions the assay responded linearly to urea concentration in the range of 0.25 to 3 mM with a correlation coefficient of 0.996. This study also demonstrated the feasibility of a fluorometric method for urea determination with fiber optic detection. The probe response also displayed good linearity in the range of 0.10 to 1.75 mM urea.

Murray, Karen R.; Nair, Maya P.; McConathy, Walter J.; Lacko, Andras G.

doi: 10.1080/00032719908542915pmid: N/A

Several immunoassay models, including sandwich ELISA, solid phase ELISA and sink immunoassay and antibody combinations were investigated to develop an ELISA assay for LCAT with an appropriate linear range and sensitivity. Solid phase immunoassays were found to be most suitable for measuring LCAT from cell culture medium and in partially purified preparations. The immunoassays were analyzed for matrix interference, recovery studies, intra-run precision and inter-run precision. These studies have identified a reliable method for measuring LCAT in purified preparations and cell culture media, and provide the foundation for further development of immunoassays for clinical application.

Ghindilis, Andrey L.; Rabinovich, Emmanuil; Lopez, Gabriel P.

doi: 10.1080/00032719908542916pmid: N/A

An immuno-displacement assay technique utilizing disposable sensing elements and based on fluorescence detection of rhodamine labeled immuno-complexes has been developed. Enkephalin has been used as a model analyte. The disposable sensing element is a strip coated with a thin gold layer by vacuum evaporation. The gold surface is modified by immobilized enkephalin. The enkephalin layer is treated with a rhodamine labeled immuno-complex against enkephalin to form a fluorescent layer. A presence of free enkephalin in the media causes a displacement of the labeled immuno-complex. This results in a decrease in surface fluorescence, which is proportional to the concentration of the free enkephalin (analyte). A detection range 1-100 μM for the enkephalin assay has been observed for a 10 min assay time.

Amin, Alaa S.

doi: 10.1080/00032719908542917pmid: N/A

Four azo compounds based on diazotization of 2-aminobenzothiazole have been synthesized and characterized by elemental analysis as well as different spectroscopic techniques. The potentiality of the prepared compounds as new chromogenic reagents for the spectrophotometric determination of Mo6+ was studied by extensive investigation of optimum conditions favouring the formation of the coloured complexes. Beers law is obeyed in the concentration range 0.2-8.5 μg ml−1 whereas Ringbom optimum concentration range was 0.8-7.5 μg ml−1. The detection limit was 0.05 μg ml−1. The molar absorptivity and Sandell sensitivity of the formed complexes are calculated. The effect of interfering ions on the determination of Mo6+ was investigated. The relative standard deviations for six replicate determinations of 5.0 μg ml−1 of Mo6+ are 1.23, 1.47, 1.05 and 1.38 % using reagents I, II, III and IV, respectively. The proposed method has been applied to investigate the amount of Mo6+ in human urine samples. The molybdenum levels found between 0.5-2.1 μg/100 ml.

Kudo, Masakiyo; Ohkubo, Tadashi; Sugawara, Kazunobu

doi: 10.1080/00032719908542918pmid: N/A

A high-performance liquid chromatography (HPLC) assay was developed for the determination of fleroxacin in plasma. The plasma samples were directly introduced onto a HPLC column after filtering through a MolcutII® membrane filter, which removes high molecular weight proteins. The fleroxacin in filtrate was separated from interfering substances and retained on a pre-column using an ODS stationary phase and then was introduced to an analytical column with an ODS stationary phase by column switching. Fleroxacin and lomefloxacin, as an internal standard, were detected by ultraviolet absorbance at 295 nm. Determination of fleroxacin was possible over the concentration range 50-4000 ng/ml; the limit of detection was 20 ng/ml. The recovery of fleroxacin added to plasma was 97.3-100.4% with a coefficient of variation of less than 2.2%. This method is applicable to drug level monitoring in the plasma of patients being treated with fleroxacin and of healthy volunteers participating in pharmacokinetic studies.

Radi, A.; EL Ries, M. A.; Bekhiet, G. E.

doi: 10.1080/00032719908542919pmid: N/A

The electrochemical oxidation of gliclazide has been studied in Britton-Robinson buffers in the pH range of 2.0-12.0 by cyclic voltammetry and differential pulse polarography at carbon paste electrode. Gliclazide gave rise to two voltammetric peaks, corresponding to the oxidation of hydrazide –CO-NH-NR2 moiety (where R2 are cycloalkylamino ring). The oxidation process has been shown to be irreversible and diffusion-controlled with adsorption characteristics over the entire pH range. Analytical methods with adequate precision and accuracy have been developed for the voltammetric determination of gliclazide in tablets.

Kargosha, Kazem; Ahmadi, Seyyed Hamid

doi: 10.1080/00032719908542920pmid: N/A

A first-derivative FTIR spectrometric method has been developed for the simultaneous determination of sulphamethoxazole (SMX) and trimethoprim (TMP) in their binary dosage formulations. The procedure is based on extraction of active components in a solution of 30% v/v acetone in chloroform, filtration and measurement of the amplitudes of the first derivative spectrum at 1138 and 1166 cm−1 for TMP and SMX respectively. Good linearity, accuracy and precision were found for this method. The relative standard deviation for TMP and SMX were found to be 1.5 and 1.3% in a commercial Co-trimoxazole tablet, respectively (n=5).