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PubMed1998

Sensitive quantitation of chromium-DNA adducts by inductively coupled plasma mass spectrometry with a direct injection high-efficiency nebulizer.

A novel method is described for the sensitive detection of chromium-DNA adducts. Chromium-DNA adducts were determined in 1 microgram of DNA from normal human lung fibroblasts exposed to sodium chromate using microscale flow injection analysis with a direct injection high-efficiency nebulizer and inductively coupled plasma mass spectrometry detection. The frequency of Cr-DNA adducts increased in a dose-dependent sigmoidal manner, indicating saturation and toxicity. The low detection limits (on the order of parts per trillion) allows the detection of as few as two Cr adducts per 10,000 bases, which, coupled with the small DNA sample requirement, makes this technique suitable for measuring metal-DNA adducts as biomarkers of exposure to toxic and carcinogenic metals such as Cr, in cultured cells, animals, and humans.

Chromium↗

PubMed1994

Automated imunoanalysis systems for monitoring mammalian cell cultivation processes.

Two different automated immunoanalysis systems are presented. Both are based on the principles of flow-injection analysis and were developed to provide reliable, rapid monitoring of relevant proteins in animal cell cultivation processes. One system uses a turbidimetric analysis, and the other employs a heterogeneous chemistry with immobilized immunocomponents. For both systems, the analysis time is in the range of a few minutes, and a complete analysis cycle, including triplicate analyses and various washing steps, is in the range of 20-30 minutes. Samples from cultivation processes can be analyzed directly without dilution. Quantitation of proteins such as rt-PA or monoclonal antibodies can be performed over an analyte concentration range of 1-1000 mg/L. Both systems were compared to conventional ELISA assays on microtiter plates. The turbidimetric analysis system also included a biosensor for simultaneous glucose determination.

Animals↗

PubMedDate not supplied

Biosensors for process monitoring.

A short review about the biosensor research activities for bioprocess monitoring in the F.R.G. after its reunification is given. The principles of biosensor applications are presented. In situ sensors and sensors based on the principles of flow injection analysis are studied. Some applications of a four-channel enzyme thermistor, bio-field effect transistors, and immunoanalysis systems for real process monitoring are presented.

Biosensing Techniques↗

PubMed1992

Chromium (VI) reducing capacity of ascorbic acid and of human plasma in vitro.

In the metabolism of chromium(VI) its reduction in human plasma is of importance; an extracellular reduction of Cr(VI) is regarded as a detoxification step. Ascorbic acid has been suggested to represent the majority of the Cr(VI)-reducing capacity of human plasma. Therefore the kinetics of the reaction of Cr(VI) with ascorbic acid, at biologically realistic concentrations were studied. Ascorbic acid, in 0.2 M HEPES buffer and at concentrations ranging from 14.2 to 113.6 nmol ml-1 (2.5-20.0 microgram ml-1), was mixed with Cr(VI) (0.4-1.5 nmol ml-1) and incubated at pH 7.4 and 37 degrees C. In addition, chromate solutions at different concentrations [1.5-100 nmol ml-1 Cr(VI)], were incubated at 37 degrees C with freshly drawn blood. From these incubates, ascorbic acid and its oxidized form, dehydroascorbic acid, were simultaneously analyzed by HPLC and post-column derivatization. Chromate was determined by flow injection analysis. The reaction kinetics of ascorbic acid in HEPES buffer with Cr(VI) is of pseudo-first order at higher concentrations, whilst apparently at lower concentrations kinetics are consistent with an autocatalyzed reaction. Results obtained after spiking human plasma are similar. However, when Cr(VI) was reacted with human plasma, no changes in the intrinsic contents of ascorbic acid of the plasma samples occurred. Also, comparing different plasma samples the intrinsic plasma contents of ascorbic acid and the reduction capacities for Cr(VI) [ranging between 0.48 and 0.63 nmol ml-1 Cr(VI) to be reduced] did not correlate. This shows that the reduction of Cr(VI) in native human plasma is complex and is not only determined by the plasma ascorbic acid levels. This is in contrast to the situation in lung lavage fluids (Suzuki 1988; Suzuki and Fukuda 1990) where the concentrations of ascorbic acid are much higher than in blood.

Ascorbic Acid↗

PubMedDate not supplied

Improved biosensor for glucose based on glucose oxidase-immobilized silk fibroin membrane.

Based on glucose oxidase-immobilized silk fibroin membrane and oxygen electrode, the authors have developed an amperometric glucose sensor in flow-injection analysis. After the sensor was improved by the configuration of oxygen electrode and a temperature control system was added to the electrode body, its sensitivity, analytical precision, and stability were enhanced greatly. The authors first introduced a tailing inhibitor-ion pair reagent into a buffer system in the biosensor so as to eliminate all interference from hemacyte, macromolecules, and small mol wt charged species besides electroactive specie ascorbate in complex matrices. A considerably serious tailing of the biosamples, such as whole blood, plasma, serum, or urine on the sensor, based on enzyme electrode, entirely disappeared, their response times were shortened, and base lines became more smooth and stable. The glucose sensor has a broad range of linear response for glucose (up to 25.0 mmol/L) and a good correlation (gamma = 0.999) under conditions of control temperature 32.0 degrees C and 1.6 mL/min 0.02 mol/L phosphate buffer containing 0.5% tailing inhibitor (v/v). Recoveries of glucose in these biosamples are within the range of 93.71-105.88%, and its repeatabilities for determining glucose, repeated 100 times, human blood dilution 125 times, and serum 128 times, are 1.81, 2.48, and 2.91% (RSD), respectively. The correlation analysis for 200 serum samples showed that the correlation (gamma) is 0.9934 between the glucose sensor and Worthington method for determining serum glucose used conventionally in a hospital laboratory. Moreover, the enzyme membrane used in the biosensor can be stored for a long time (over 2 yr) and measured repeatedly over 1000 times for biosamples. The glucose sensor is capable of detecting over 60 biosamples/hr.

Animals↗

PubMedDate not supplied

Microfabrication and microsensors.

In order to fabricate microsensors, a micromachining, microfabrication technique based on LSI technologies has been used. In this paper, useful micromachining techniques are described. Examples of integrated microsensors, such as a pressure sensor including a readout circuit and a multisensor cell for blood gas analysis, are introduced. Microflow control devices, such as microvalve and a micropump for integrated chemical analysis systems, are also described.

Blood Gas Analysis↗

PubMed2003

The pattern of urinary acylcarnitines determined by electrospray mass spectrometry: a new tool in the diagnosis of diabetes mellitus.

l-Carnitine and its esters are products of intermediary metabolism of organisms. The distribution pattern or the favored excretion of individual acylcarnitines tells something about metabolic diseases. The determination of the urinary acylcarnitine pattern by flow injection analysis (FIA)-electrospray ionization (ESI)-mass spectrometry (MS) is presented. Groups of healthy probands and patients suffering from diabetes mellitus were investigated due to their significant acylcarnitine profile. The statistical analysis of data sets obtained clearly shows a difference in the acylcarnitine pattern of healthy and sick probands. In comparison to the controls, diabetes mellitus patients excrete more long-chain carnitine esters ranging from dodecanoyl to palmitoylcarnitine. Thus, the urinary acylcarnitine pattern determined by ESI-MS can be a useful tool in the diagnosis and therapy monitoring of diabetes mellitus.

Adult↗

PubMed2003

Sensitive flow-injection chemiluminescence determination of terbutaline sulfate based on enhancement of the luminol-permanganate reaction.

A novel and highly sensitive chemiluminescence (CL) method for the determination of terbutaline sulfate, coupled with flow-injection analysis (FIA), is described in this paper. The method is based on enhancement by terbutaline sulfate of the chemiluminescence emission of the luminol-permanganate system under alkaline conditions. Under the conditions selected the concentration of terbutaline sulfate is proportional to CL intensity in the range 5 x 10(-10)-5 x 10(-7) g mL(-1), with a detection limit of 1.7 x 10(-10) g mL(-1) (3 sigma). The relative standard deviation is 2.8% for 1 x 10(-8) g mL(-1) terbutaline sulfate ( n=11). Ninety samples can be determined per hour. The proposed method has been used to determine terbutaline sulfate in pharmaceutical preparations and in plasma and urine samples with satisfactory results. The possible mechanism of the chemiluminescence reaction is discussed briefly.

Calibration↗

PubMed2004

Determination of trace amounts of phosphate by flow-injection photometry.

This paper describes a simple and rapid procedure for determination of traces of phosphate by means of molybdenum blue chemistry. The use of a cost-effective home-made flow cell with a long path length in combination with a light emitting diode (LED) and a photodiode (PD) is demonstrated as a simple absorbance detector for flow-injection analysis. In this method, a sample is injected into the carrier stream through an injection valve and mixed online with mixed reagent (a mixture of molybdate, bismuth, and ascorbic acid in sulfuric acid). The color intensity of the resulting association complex, molybdenum blue, is measured photometrically ( lambda(max) 875 nm). The proposed method can be used to detect phosphate in the range 0.02-4.0 mg x L(-1) and the precision of the proposed procedure is less than 5% at 0.1 mg x L(-1) phosphorus as phosphate. The method has been successfully applied to a variety of natural water samples.

Journal Article↗

PubMed2005

Development of a passive micromixer based on repeated fluid twisting and flattening, and its application to DNA purification.

We have developed a three-dimensional passive micromixer based on new mixing principles, fluid twisting and flattening. This micromixer is constructed by repeating two microchannel segments, a "main channel" and a "flattened channel", which are very different in size and are arranged perpendicularly. At the intersection of these segments the fluid inside the micromixer is twisted and then, in the flattened channel, the diffusion length is greatly reduced, achieving high mixing efficiency. Several types of micromixer were fabricated and the effect of microchannel geometry on mixing performance was evaluated. We also integrated this micromixer with a miniaturized DNA purification device, in which the concentration of the buffer solution could be rapidly changed, to perform DNA purification based on solid-phase extraction.

Chemical Fractionation↗

PubMed2005

Establishment of signal-recovery functions for calculation of recovery factor. Application to monitoring of contaminant residues in vegetables by chemiluminescence detection.

A new strategy is proposed for verifying if recovery factor is constant and independent of the real analyte content of samples. A signal-recovery function has been developed on the basis of measurement of spiked test samples before and after a pre-treatment step and considering, as starting point, a recent IUPAC recommendation which distinguishes between two terms--recovery factor, R, and apparent recovery, R*. Apparent recovery includes recovery factor and a new recovery term proposed in a previous paper by the authors, named calibration recovery, R(C). The signal-recovery function is obtained directly from the measured analytical signals instead of from the concentrations, simplifying the calculations. A linear signal-recovery curve indicates that the recovery factor is constant in the analyte concentration range studied experimentally and, in this way, a single recovery factor can be calculated. The usefulness of the proposed method has been shown by quantification of the pesticide carbaryl by two different flow-injection analysis methods with chemiluminescent detection based on the luminol and TCPO systems. Good results were obtained from both methods.

Food Contamination↗

PubMed2005

Arsenic speciation in xylem sap of cucumber (Cucumis sativus L.).

Flow injection analysis (FIA) and high-performance liquid chromatography double-focusing sector field inductively coupled plasma mass spectrometry (HPLC-DF-ICP-MS) were used for total arsenic determination and arsenic speciation of xylem sap of cucumber plants (Cucumis sativus L.) grown in hydroponics containing 2 micromol dm(-3) arsenate or arsenite, respectively. Arsenite [As(III)], arsenate [As(V)] and dimethylarsinic acid (DMA) were identified in the sap of the plants. Arsenite was the predominant arsenic species in the xylem saps regardless of the type of arsenic treatment, and the following concentration order was determined: As(III) > As(V) > DMA. The amount of total As, calculated taking into consideration the mass of xylem sap collected, was almost equal for both treatments. Arsenite was taken up more easily by cucumber than arsenate. Partial oxidation of arsenite to arsenate (<10% in 48 h) was observed in the case of arsenite-containing nutrient solutions, which may explain the detection of arsenate in the saps of plants treated with arsenite.

Arsenates↗

PubMed2006

Flow screen-printed amperometric detection of p-nitrophenol in alkaline phosphatase-based assays.

p-Nitrophenyl phosphate is one of the most widely used substrates for alkaline phosphatase in ELISAs because its yellow, water-soluble product, p-nitrophenol, absorbs strongly at 405 nm. p-Nitrophenol is also electroactive; an oxidative peak at 0.97 V (vs. an Ag pseudoreference electrode) is obtained when a bare screen-printed carbon electrode is used. When an amperometric detector was coupled to a flow-injection analysis system the detection limit achieved for p-nitrophenol was 2x10(-8) mol L(-1), almost two orders of magnitude lower than that obtained by measuring the absorbance of the compound. By use of this electrochemical detection method, measurement of 7x10(-14) mol L(-1) alkaline phosphatase was achieved after incubation for 20 min. The feasibility of coupling immunoassay to screen-printed carbon electrode amperometric detection has been demonstrated by performing an ELISA for detection of pneumolysin, a toxin produced by Streptococcus pneumoniae, which causes respiratory infections. The method is simple, reproducible, and much more sensitive than traditional spectrophotometry.

Alkaline Phosphatase↗

PubMed2007

Development of a novel nitrate-selective composite sensor based on doped polypyrrole.

The manufacture and evaluation of a novel sensor built with a composite material, highly selective to nitrate ions using doped polypyrrole as a recognition agent, are presented. When the ratio of recognition agent to graphite was optimized at 1:1, and the sensitivities found closely approached nernstian behavior. The stability times attained were less than 14 min with response times also below 20 s. Batch characterization of the sensor displayed a sensitivity of 57.1 mV/decade of nitrate ion activity (alphaNO3-) and a detection limit of 5.37 x 10(-5) M, which are comparable to those reported for commercial sensors. Evaluation of the selectivity coefficients showed high affinity to nitrate ion, superior to that of commercial sensors and others reported in the literature. The composite material gives the sensor a prolonged service life with the added capability of allowing the regeneration of its active surface. Coupling the sensor and a solid state, composite-type, reference electrode to a flow injection analysis system (FIA) permitted to achieve an effective overall assessment of the system. A nitrate determination test was conducted on real samples. A comparison of the results obtained, either with stationary measurements or with FIA, indicated that there were no significant differences from the values from manufacturer's specifications.

Calibration↗

PubMed2000

Flow injection spectrophotometric determination of ultra trace amounts of oxalic acid.

A new simple, sensitive and rapid catalytic-spectrophotometric method for the determination of oxalic acid has been described based on its catalytic effect on the redox reaction between dichromate and Brilliant cresyl blue in acidic media by means of a flow injection analysis method. The color change of Brilliant cresyl blue due to its oxidation was monitored spectrophotometrically at 625 nm. The calibration graph was linear in the range of 0.020-4.70 microg/mL oxalic acid with a limit of detection 0.005 microg/mL of oxalic acid. The relative standard deviation for ten replicate measurements of 0.020 microg/mL and 0.900 microg/mL was 2.2% and 1.7%, respectively. No serious interference was identified. Oxalic acid was determined in wastewater and in spinach by the proposed method with satisfactory results.

Hydrogen-Ion Concentration↗

PubMed2000

Automized procedures for the determination of ozone and ammonia contents in air by using the chromatomembrane method for gas-liquid extraction.

Chromatomembrane cells (CMC) operate as unique manifolds for extraction and preconcentration procedures in computer controlled flow-injection-analysis (FIA). By coupling to an ion chromatograph and to a conductometer, respectively, instrumentation is obtained that allows the pretreatment and the detection of ozone- and ammonia-containing samples after absorption of the gaseous constituents of air inside a chromatomembrane cell. The analysis of air even in the presence of liquid aerosols is discussed.

Air Pollution↗

PubMedDate not supplied

Bio- and chemiluminescence in bioanalysis.

Analytical chemiluminescence and bioluminescence represent a versatile, ultrasensitive tool with a wide range of applications in diverse fields such as biotechnology, pharmacology, molecular biology, clinical and environmental chemistry. Enzyme activities and enzyme substrates and inhibitors can be efficiently determined when directly involved in luminescent reactions, and also when they take part in a reaction suitable for coupling to a final light-emitting reaction. Chemiluminescence detection has been exploited in the fields of flow-injection analysis and column-liquid chromatographic and capillary-electrophoretic separative systems, due to its high sensitivity when compared with colorimetric detection. It has widely been used as an indicator of reactive oxygen species formation in cells and whole organs, thus allowing the study of a number of pathophysiological conditions related to oxidative stress. Chemiluminescence represents a sensitive and rapid alternative to radioactivity as a detection principle in immunoassays for the determination of a wide range of molecules (hormones, food additives, environmental pollutants) and in filter membrane biospecific reactions (Southern, Northern, Western, dot blot) for the determination of nucleic acids and proteins. Chemiluminescence has also been used for the sensitive and specific localization and quantitation of target analytes in tissue sections and single cells by immunohistochemistry and in situ hybridization techniques. A relatively recent application regards the use of luminescent reporter genes for the development of bioassays based on genetically engineered microorganisms or mammalian cells able to emit visible light in response to specific inorganic and organic compounds. Finally, the high detectability and rapidity of bio- and chemiluminescent detection make it suitable for the development of microarray-based high throughput screening assays, in which simultaneous, multianalyte detection is performed on multiple samples.

Animals↗

PubMed2000

Potentiometric determination of acids and bases using a silica gel based carbon-epoxy indicator electrode.

The construction and the application of a silica gel based carbon-epoxy indicator electrode for the potentiometric determination of acids and bases are described. The effect of composition of silica gel and carbon-epoxy, slope (mV/pH), linear response (pH range) and the use for acid-base titrations were investigated. The data obtained for the acid-base titrations were compared with those obtained using a glass electrode in the same conditions. The electrode showed a linear response in the pH 2 to 13 range with a slope of -40.5 +/- 0.4 mV/pH (at 25 degrees C) and a response time of less than 15 s. The lifetime of the electrode was higher than one year (over 6000 determinations) with a decrease of only 5% of the initial potentiometric response. The silica gel based carbon-epoxy electrode showed excellent results in the end-point indication potentiometric titrations in determination of acids and bases. The miniaturization of the proposed electrode for flow injection analysis was investigated.

Journal Article↗