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Results for “Flow Injection Analysis”

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PubMed1988

Determination of glutathione in biological material by flow-injection analysis using an enzymatic recycling reaction.

A sensitive and specific assay for glutathione using a recycling reaction followed by spectrophotometric detection in a flow-injection analysis system is presented. The proposed method provides specific amplification of the response to glutathione by combined use of the enzyme GSSG reductase and the chromogenic reagent 5,5'-dithiobis(2-nitrobenzoic acid). Both oxidized (GSSG) and reduced (GSH) glutathione are detected, so that GSSG must be determined separately after alkylation of the GSH with N-ethylmaleimide. The sensitivity is controlled by the number of times the cycle occurs and therefore by the residence time of the sample in the reactor. This time depends on the reactor length and the flow rate. The influence of residence time, temperature, and enzyme concentration on the response has been studied and the optimum reaction conditions have been selected. The sample throughput is as high as 30 h(-1) and the detection limit is 1 pmol GSH at a signal-to-noise ratio of 3. The method has been evaluated by the quantification of GSH and GSSG in isolated hepatocytes. A high correlation between the new flow-injection analysis method and the original spectrophotometric batch assay has been found (slope = 1.039, intercept = 0.6, n = 216, r = 0.977). The main advantages of the proposed method are high sample throughout, high sensitivity, and good reproducibility.

Animals↗

PubMedDate not supplied

Determination of sulfite in food by flow injection analysis.

A method is described for the determination of sulfite levels in food products by flow injection analysis (FIA). The method is based on the decolorization of malachite green by SO2, which is isolated from the flowing sample stream by means of a gas diffusion cell. The FIA method has a detection limit in food sample extracts of 0.1 ppm SO2 (3 times peak height of blank), which corresponds to 1-10 ppm SO2 in a food product, depending on the extraction procedure used. At the 5 ppm SO2 level in a food extract, the precision of replicate injections is +/- 1-2%. The method was tested on a variety of both sulfite-treated and untreated food products and the results compared favorably with those obtained by the Monier-Williams, colorimetric (pararosaniline), and enzymatic (sulfite oxidase) methods. The average differences from the FIA results were 19, 11, and 12%, respectively, for those samples (n = 12) above 50 ppm SO2. At lower levels the results were somewhat more erratic due to inaccuracies of the various methods at low concentrations.

Colorimetry↗

PubMedDate not supplied

Determination of free (pH 2.2) sulfite in wines by flow injection analysis: collaborative study.

A method for the determination of free sulfite in wine by flow injection analysis (FIA) is described. The method involves liberation of sulfur dioxide from the wine at pH 2.2, with detection by decolorization of a malachite green solution. The method was collaboratively studied, and the results indicated an average reproducibility of 12% for white wine samples (average level 12.1 ppm SO2) and 26% for red wine samples (average level 3.1 ppm). When the FIA method was compared to an aeration/oxidation method, the results indicated a high degree of correlation between the 2 methods. The FIA method has been adopted by AOAC official first action.

Hydrogen-Ion Concentration↗

PubMed1992

Amperometric assay of creatinine in urine by flow injection analysis based on conjugated reactions of immobilized enzymes. Simultaneous compensation of endogenous ammonia.

A flow-injection analysis biosensor system was developed for the amperometric assay of creatinine based on coupled reactions of three immobilized enzymes, using an oxygen electrode as the detection device. The ammonia produced by creatinine deiminase-catalyzed hydrolysis of creatinine was further converted into L-glutamate with two sequentially aligned enzyme reactors: glutamate dehydrogenase and glutamate oxidase. Endogenous ammonia was simultaneously compensated with a double peak recording system, where the flow was split after sample injection and rejoined before the glutamate dehydrogenase reactor. The system gave linear calibration in a range of 0.1-2.0 mM for creatinine and the first peak of ammonia, and 0.1-3.0 mM for the second peak of ammonia. One run was completed within two minutes. The system can be readily applied to the assay of creatinine in urine and showed good correlation with that from the currently used Jaffe method.

Amino Acid Oxidoreductases↗

PubMed1979

Determination of total CO2 in plasma by automated flow-injection analysis.

We describe a procedure for measuring total CO2 in plasma, based on the principles of the flow-injection analysis technique, which makes use of unsegmented fast-flowing reagent streams, as developed by Růziĉka and co-workers. The further methodological design resembles the silicone-rubber membrane technique of Kenny and Cheng. CO2 in the sample is released by reaction with H2SO4. Appropriate amounts of CO2 permeate through the membrane that separates the acid reagent streams and a buffered cresol-red indicator system. The experimental set-up and functioning of this system are described.

Autoanalysis↗

PubMed1990

Use of ionomer membranes to enhance the selectivity of electrode-based biosensors in flow-injection analysis.

The use of ionomer membranes to enhance the selectivity of potentiometric enzyme electrodes in flow-injection measurement arrangements is examined. The ionomer membranes employed are permeable to analyte substrates but relatively impermeable to detectable ions that would normally interfere with the measurement of the substrates if the enzyme electrodes were in direct contact with the sample. As a model system, the selectivity of enzyme electrodes prepared with nonactin-based ammonium-sensitive polymeric membranes is evaluated. In the preferred configuration, a thin hydrophilic anion-exchange membrane is incorporated within a flow-through dialysis unit upstream from the enzyme-electrode detector. As the sample passes through the dialysis unit, neutral or anionic analyte molecules (urea or glutamine) move through the membrane while the permeation of endogenous ammonium ions and other cations in the sample is retarded. A flowing recipient buffer on the other side of the membrane carries the analyte substrate to the enzyme-electrode detector. Enhancements in selectivity for analyte substrates over endogenous ammonium and potassium ions are greater than or equal to 9-fold when compared to enzyme-electrode flow-injection analysis (FIA) systems assembled without the ionomer membrane unit. The analytical utility of the proposed system is demonstrated by the accurate measurements of urea in blood serum and L-glutamine in hybridoma bioreactor media.

Biosensing Techniques↗

PubMed1992

Determination of urinary glucose by a flow injection analysis amperometric biosensor and ion-exchange chromatography.

A practical biosensor system has been developed for the determination of urinary glucose using a flow-injection analysis (FIA) amperometric detector and ion-exchange chromatography. Glucose oxidase was immobilized onto porous aminopropyl glass beads via glutaraldehyde activation to form an immobilized enzyme column. On the basis of its negative charge at pH 5.5, endogenous urate in urine samples was effectively retained by an upstream anion-exchange resin column. The biosensor system possessed a sensitivity of 160 +/- 2.4 RU microM-1 (RU or relative unit is defined as 2.86 microV at the detection output) for glucose with a minimum detection level of 10 microM. When applied for the determination of urinary glucose, the result obtained compared very well with that of the widely accepted hexokinase assay. The immobilized glucose oxidase could be reused for more than 1000 repeated analyses without losing its original activity. The reuse of the acetate anion-exchange column before replacement would be about 25-30 analyses. Acetaminophen and ascorbic acid were also effectively adsorbed by the acetate anion exchanger. The introduction of this type of anion exchanger thus greatly improved the selectivity of the FIA biosensor system and fostered its applicability for the determination of glucose in urine samples.

Adult↗

PubMed1992

Flow-injection analysis for malondialdehyde in plasma with the thiobarbituric acid reaction.

A simple, precise, and rapid method to measure plasma malondialdehyde (MDA) was developed by use of solvent extraction--flow-injection analysis. The reagent solution, containing thiobarbituric acid (TBA), 5 g/L in 100 mL/L phosphoric acid, and extraction solvent (methylisobutyl ketone, MIBK) were propelled with a double-plunger micropump at a flow rate of 0.3 mL/min, and 20 microL of sample was introduced into the reagent stream. After TBA-MDA reactant was extracted into MIBK, the organic phase was continuously separated by a successive phase-separation system equipped with two phase separators, and the absorbance of the TBA-MDA reactant was measured at 532 nm. This approach resulted in excellent sensitivity, a CV of < 1.5%, a good correlation with the conventional manual method, and a sampling frequency of 7 samples/h, suggesting that this semiautomated method is suitable for measuring plasma MDA.

Animals↗

PubMed1992

Colorimetric determination of free and total cholesterol by flow injection analysis with a fiber optic detector.

A flow injection method for the determination of total and free cholesterol is presented. Cholesterol esterase and cholesterol oxidase are immobilized on aminoalkyl glass beads. The beads are packed into a tubular glass reactor. The cholesterol esters traversing through the esterase reactor are cleaved to cholesterol and fatty acids. The oxidase reactor converts cholesterol to cholest-4-en-3-one and hydrogen peroxide is generated. The sample stream is merged with reagent streams consisting of a peroxidase solution and a solution of 2,2'-azino-bis-(3-ethyl-benzthiazoline-6-sulfonic acid) diammonium salt, and a hydrogen peroxide-dependent color reaction takes place in a short coiled reactor. The signal is monitored by means of fiber optic instrumentation. Cholesterol concentration can be related to the absorption of the oxidized dye form at a wavelength of 425 nm. The working range is 0.5-0.8 mmol l-1, and the sample throughputs are 60 and 30 h-1 for free and total cholesterol, respectively.

Cholesterol↗

PubMed1992

Flow injection analysis and in-line biosensors for bioprocess control: a comparison.

Miniaturization will unify the different approaches chosen for the application of biosensors in bioprocess control. The most versatile system, which in our opinion is flow injection analysis will be the method of choice for the introduction of biosensors in bioprocess control. A lot of experience will be gained for the future development of miniaturized total chemical analysis systems.

Biosensing Techniques↗

PubMed1992

Use of various types of column reactors for flow-injection analysis.

Two or three different kinds of immobilized enzymes can be aligned in a minireactor so that sequential enzymatic reactions are carried out from upstream to downstream during flow-injection analysis. A lactate oxidase-catalase reactor, used as precolumn for removing pre-existing lactate in serum before the lactose dehydrogenase (LDH) reactions, was useful for the determination of serum LDH activity, which did not require any blank correction. A sequential glutamate dehydrogenase-glutamate oxidase reactor was also useful for a novel chemiluminometric determination of ammonia. On the other hand, a co-immobilized creatininase-creatinase-sarcosine oxidase reactor, in spite of containing creatininase which catalyses the reversible reaction, was the most efficient for the determination of serum creatinine.

Ammonia↗

PubMedDate not supplied

Determination of total sulfite in shrimp, potatoes, dried pineapple, and white wine by flow injection analysis: collaborative study.

A method for the determination of total sulfite in shrimp, potatoes, dried pineapple, and white wine by flow injection analysis (FIA) was collaboratively studied by 8 laboratories. In the method, the sample solution is reacted with sodium hydroxide to liberate aldehyde-bound sulfite. The sample stream is acidified to produce SO2 gas, which diffuses across a Teflon membrane in the gas diffusion cell into a flowing stream of malachite green. The degree of discoloration of the malachite green is proportional to the amount of sulfite in the sample solution. Red wine was included in the study but interlaboratory precision for these samples was not satisfactory and correlation with Monier-Williams results was poor. The present method is not recommended for use with these samples. For shrimp, potatoes, dried pineapple, and white wine, average reproducibility (RSDR) of results was 25% for samples at 10 ppm SO2 and 10% for samples at greater than 50 ppm. Overall average reproducibility was 14%. Recoveries of sulfite added to samples averaged 80%. Comparison of FIA with the Monier-Williams method indicated comparable results by the 2 methods. The FIA method has been adopted official first action for determination of greater than or equal to 5 ppm total sulfite in shrimp, potatoes, dried pineapple, and white wine.

Animals↗

PubMed1991

Electrochemical enzyme immunoassay for phenytoin by flow-injection analysis incorporating a redox coupling agent.

Using phenytoin as a model analyte, we demonstrate an electrochemical enzyme immunoassay based on flow-injection analysis and incorporating 2,6-dichloroindophenol (DCIP) as a redox coupling agent. DCIP reacts with NADH to form NAD+ and DCIPH2, the reduced form of the coupling agent. The production of DCIPH2 is monitored at +250 mV vs Ag/AgCl. This low applied potential improves selectivity in the biological matrix, differentiating against components that are oxidizable at the more-positive potentials required for direct electrochemical detection of NADH. The kinetics-based assay also eliminates other common interferences, mainly from ascorbic acid and glutathione. This system does not require precolumns or analytical columns for isolation of the NADH response. Good agreement with a routine clinical laboratory procedure for phenytoin is obtained for clinical samples (r = 0.95), illustrating the feasibility of such an approach.

2,6-Dichloroindophenol↗

PubMed1990

Copper determination in urine by flow injection analysis with electrochemical detection at platinum disk microelectrodes of various radii.

The incorporation of platinum disk microelectrodes of various radii (2.5-50 microns) in a well-jet flow cell offers reduced limits of detection for the determination of copper in urine by flow injection analysis compared with standard methods based on a conventional sized glassy carbon disk macroelectrode (radius 1.5 mm), in a thin-layer cell. The radius of the platinum disk microelectrode was found to be critical with respect to both the limit of detection and flow rate dependence. An optimal radius value of 28 microns was found with detection limits increasing with both larger and smaller electrode radii. In contrast, as theoretically expected, a diminished flow rate dependence was observed the smaller the radii of the platinum disk microelectrodes. Sample cleanup and preparation is conveniently achieved by the use of Sep-Pak cartridges and formation of a copper dithiocarbamate complex. The metal complex is easily oxidized at platinum disk microelectrodes in acetonitrile, which was the solvent used in the flow injection method of analysis.

Copper↗

PubMed1988

Immobilization of linamarase and its use in the determination of bound cyanide in cassava using flow injection analysis.

Extracts from the tubers (cortex and parenchyma) and leaves of Manihot esculenta Crantz (cassava) were analyzed for their releasable cyanide content using flow injection analysis incorporating an immobilized linamarase bioreactor. Linamarase was immobilized under very mild conditions to an activated 2-fluoro-N-methylpyridinium Fractogel support. The released cyanide, which was monitored spectrophotometrically at 525 nm using an alkaline picrate reagent, was found to be highest in the cortex and lowest in the parenchyma.

Calibration↗

PubMed1992

[Construction and application of all-solid-state aconitine electrochemical detector in flow injection analysis].

A new kind of all-solid-state electrochemical detector for very toxic alkaloids such as aconitine, mesaconitine and hypaconitine has been studied. It exhibits Nernstian response for these alkaloids with a slope of 56 mV/decade over the concentration range of 3 x 10(-5)-1 x 10(-2) mol/L at pH 2-7 under the flow condition. Direct potentiometry for the determination of aconitine in Aconitum kusnezoffii Reichb., Aconitum carmichaeli Debx. and Xiaohuoluo Wan showed average recoveries of 98.5, 98.3 and 96.8% and relative standard deviations of 1.8, 2.4 and 3.5%, respectively. It can be used for the determination of very toxic alkaloids in the above mentioned samples by flow injection analysis. It also can be used for the study of the hydrolytic kinetics of aconitine.

Aconitine↗

PubMed1990

Flow injection analysis and real-time detection of RNA bases by surface-enhanced Raman spectroscopy.

Surface-enhanced Raman scattering (SERS) spectroscopy has been successfully interfaced with a flow injection analysis system to detect RNA bases in real time. Four of the major base components of RNA, uracil, cytosine, adenine, and guanine, were introduced into the flow injection system and were mixed with a Ag sol prior to SERS measurements. Several experimental parameters including pH, temperature, flow rate, and tubing materials were examined, and their impact on the SERS spectra is presented here. The feasibility of interfacing flow injection based SERS detection methods with liquid or high-performance liquid chromatography for the detection of individual components in a complex mixture is also assessed.

Chromatography, High Pressure Liquid↗

PubMed1991

Determination of proteolytic enzymes by flow-injection analysis.

Quantitation of proteolytic enzymes using N-succinyl-L-Ala-L-Ala-L-Pro-L-Phe-p-nitroanilide has been adapted to flow-injection analysis. This procedure has been developed using two different proteases: subtilisin and chymotrypsin. For both enzymes the influence of substrate concentration on spectrophotometric response has been studied. The assay is based on the merging zones technique combined with a washing step. Results are obtained in less than 15 s and samples may be run at a rate of 90/h with good reproducibility. A linear relation between peak heights and enzyme concentrations was observed for 0-0.15 Anson unit/liter of subtilisin and for 0-30 mg/liter of a commercial preparation of chymotrypsin. The method requires only small sample volumes, and the consumption of the chromogenic substrate is reduced to a minimum by using intermittent pumping.

Amino Acid Sequence↗