“Reagentless” Flow Injection Determination of Ammonia and Urea Using Membrane Separation and Solid Phase Basification

Flow injection analysis instrumentation and methodology for the determination of ammonia and ammonium ions in an aqueous solution are described. Using in-line solid phase basification beds containing crystalline media, the speciation of ammoniacal nitrogen is shifted toward the un-ionized form, which diffuses in the gas phase across a hydrophobic microporous hollow fiber membrane into a pure-water-containing analytical stream. The two streams flow in a countercurrent configuration on opposite sides of the membrane. The neutral pH of the analytical stream promotes the formation of ammonium cations, which are detected using specific conductance. The methodology provides a lower limit of detection of 10 μg/L and a dynamic concentration range spanning three orders of magnitude using a 315-μL sample injection volume. Using immobilized urease to enzymatically promote the hydrolysis of urea to produce ammonia and carbon dioxide, the technique has been extended to the determination of urea.     

Determination of urea in serum by using naturally immobilized urease in a flow injection conductimetric system

A flow injection method was developed, aimed at the determination of urea in human serum. The system makes use of the naturally immobilized urease present in Canavalia ensiformis DC (jack bean). A column is filled with small pieces of this bean, and the sample (50 microliters) containing urea passes through it carried by a 1% NaCl solution. On leaving the column the stream is merged with an alkaline reagent (0.5 mol dm-3 NaOH; 0.5% disodium dihydrogen ethylenediaminetetraacetate). The ammonium ions, arising from the enzymatic reaction that occurs inside the column, are changed into the molecular form, which permeates a polytetrafluoroethylene membrane and is received in a de-ionized water acceptor stream. The ammonia ionizes causing an increase in the conductance, which is proportional to the urea content of the sample. About 40 samples can be processed in 1 h with negligible carry-over and with a relative standard deviation of 1% or less. The results are in agreement with those obtained by a standard spectrophotometric method.     

Sequential flow injection analysis of ammonium and nitrate using gas phase molecular absorption spectrometry

A flow injection method on the basis of gas phase molecular absorption is described for the sequential determination of ammonium and nitrate. Two hundred microliters of sample solution is injected into the flow line. For ammonium determination, the sample zone is directed to a line in which reacts with NaOH (13 M) and produces ammonia. But for nitrate determination, the sample zone is passed through the on-line copperized zinc (Zn/Cu) reduction column and produces ammonium ion and in the follows ammonia. The produced ammonia in both cases is purged into the stream of N₂ carrier gas. The gaseous phase is separated from the liquid phase using a gas-liquid separator and then is swept into a flow through cell, which has been positioned in the cell compartment of an UV-Vis spectrophotometer. The absorbance of the gaseous phase is measured at 194 nm. Under selected conditions for sequential analysis of ammonium and nitrate, linear relations were found between the peak heights of absorption signals and concentrations of ammonium (10-650 μg ml⁻¹) and nitrate (20-800 μg ml⁻¹). The limit of detections for ammonium and nitrate analysis were 8 and 10 μg ml⁻¹, respectively. The relative standard deviations of repeated measurements of 50 μg ml⁻¹ of ammonium and nitrate were 2.0, 2.9%, respectively. Maximum sampling rate was about 40 samples/h. The method was applied to the determination of ammonium in pharmaceutical products and the sequential determination of ammonium and nitrate in spiked water samples.     

Sequential determination of urea and HMTA in the process solution of sol-gel route of advanced nuclear fuel fabrication

Determinations of hexamethylene tetramine (HMTA) and urea in the process solutions are required to optimize their concentrations for obtaining high quality ceramic oxide microspheres, for monitoring the washing procedure and for their subsequent recovery, recycling or waste disposal. Determination of urea in the feed solution by conventional procedures is difficult as it contains HMTA. It is more so in the effluent as it contains hydrolytic products like formaldehyde, methylol derivatives of urea, ammonium nitrate and ammonium hydroxide used for washing the gel microspheres. This work describes a derivative potentiometric method using a microprocessor-based autotitrator. Peaks on the first derivative of the titration plot corresponded to constituents of different basicities. Urea was selectively hydrolyzed at room temperature by the catalytic action of urease enzyme leaving HMTA unaffected. Ammonium hydroxide and ammonium bicarbonate produced from urea and HMTA were sequentially titrated for the analysis of the feed solution to obtain the three corresponding peaks respectively. Two separate titrations were required for the analysis of the effluent solution, which contained free ammonia also. One aliquot was first titrated directly without adding urease (for free ammonia and HMTA) and another aliquot was titrated after treatment with urease. The end points due to the ammonia used for washing and that from urea hydrolysis merged resulting in the appearance of three peaks again. Using this sequential method the relative standard deviations were found to be 0.81% and 1.38% for urea and HMTA, respectively, in eight determinations when the aliquots contained 50 to 75 mg of urea and 75 to 125 mg of HMTA. Feed and effluent solutions of the process stream were analyzed.     

A membraneless gas-diffusion unit-multisyringe flow injection spectrophotometric method for ammonium determination in untreated environmental samples

A new design of a membraneless gas-diffusion (MGD) unit coupled to a multisyringe flow injection system is proposed. The spectrophotometric determination of ammonium using an acid-base indicator was chosen to show the feasibility of this approach. Hence, in alkaline medium, ammonium ions are transformed into ammonia (donor channel) which diffuses through the headspace into the acceptor stream (bromothymol blue solution), causing a pH change and subsequently a colour change. The exploitation of the enhanced potentialities of this re-designed MGD device was the main purpose of the present work. Hence, several strategies concerning flow management were studied seeking to characterize and improve the analytical features of the methodology and moreover, untreated environmental samples were analysed without previous filtration. Consequently, stopped flow in acceptor channel with continuous flow in donor channel was chosen for the application to wastewater and spiked river water samples. A linear concentration range between 10.0 and 50.0 mg L⁻¹ of NH₄⁺, a limit of detection of 2.20 mg L⁻¹ and a determination frequency of 11 h⁻¹ were obtained.     

Simultaneous determination of ammonia and urea with an asymmetric merging zones/flow-injection configuration

A method is proposed for the simultaneous determination of urea and ammonia using a reagent-injection configuration that includes a dual injection valve (for insertion of Nessler's reagent and for accommodating the enzyme reactor). The resolution of the two peaks obtained on each injection allows the determination of both analytes in mixtures. The determination range is 1–5 μg ml^?1 for ammonia and 1–6 μg ml^?1 for urea, with relative standard deviations of 1.13% and 2.31% for ammonia (first and second peaks) and 1.86% for urea.     

A multi-commuted flow injection system with a multi-channel propulsion unit placed before detection: Spectrophotometric determination of ammonium

A flow system with a multi-channel peristaltic pump placed before the solenoid valves is proposed to overcome some limitations attributed to multi-commuted flow injection systems: the negative pressure can lead to the formation of unwanted air bubbles and limits the use of devices for separation processes (gas diffusion, dialysis or ion-exchange). The proposed approach was applied to the colorimetric determination of ammonium nitrogen. In alkaline medium, ammonium is converted into ammonia, which diffuses over the membrane, causing a pH change and subsequently a colour change in the acceptor stream (bromothymol blue solution). The system allowed the re-circulation of the acceptor solution and was applied to ammonium determination in surface and tap water, providing relative standard deviations lower than 1.5%. A stopped flow approach in the acceptor stream was adopted to attain a low quantification limit (42 μg L⁻¹) and a linear dynamic range of 50–1000 μg L⁻¹ with a determination rate of 20 h⁻¹.     

Preparation and Response Properties of Selective Bio-Electrodes Utilizing Polymer Membrane Electrode-Based Ammonia Gas Sensors

Abstract

A new type of potentiometric ammonia gas sensor is employed in the preparation of selective bio-electrodes for urea and glutamine. The bio-electrodes are constructed by immobilizing the enzyme urease and intact porcine kidney cells, respectively, at the surface of a disposable ammonium selective polymer membrane electrode-based ammonia gas sensor. The resulting electrodes have favorable response properties when compared to corresponding devices previously assembled with costly commercial gas sensors. Preliminary studies with the urea electrode demonstrate its usefulness for the rapid determination of urea in serum samples.     

Determination of total nitrogen in food by flow injection analysis (FIA) with a potentiometric differential detection system

A flow injection set-up based on potentiometric detection and gas diffusion device for the determination of total nitrogen in food is described. The detection system consisted of two ammonium-sensitive electrodes placed sequentially and each alternately operating as reference electrode. Tubular electrodes without an inner reference solution were prepared with a PVC membrane composed of nonactin in Tris (2-ethylhexyl) phosphate and potassium tetrakis (4-chlorophenyl) borate to reduce the membrane resistance. The food sample digests were inserted into the system, and the ammonium present was converted into ammonia gas. The gas diffused through a gas-permeable membrane to a buffer acceptor stream with a pH that ensured transformation to the ammonium cation, which was potentiometrically detected. Good agreement between FIA results and those provided by the reference procedure was obtained, with relative deviation errors below 5%. Using the proposed system, low reagent consumption is possible, a sampling rate of about 30 samples/h was achieved, as well as a good reproducibility for consecutive injections of the same sample (variation coefficient < 2%).     

Gas-diffusion separation and flow injection potentiometry

Summary The similarities and differences of the operation principle of gas-sensing electrodes and potentiometric detection coupled to gas-diffusion separation in flow injection analysis are discussed with special emphasis on selectivity and sensitivity aspects. Several examples of application are presented highlighting the improvements in detectability obtained by gas-diffusion flow injection potentiometry. High sensitivity determination of ammonium is achieved through accumulation of ammonia released from the sample stream in the small recipient volume of the gas-diffusion unit. A method for almost specific determination of cyanide is presented making use of gas-diffusion separation of hydrogen cyanide and potentiometric detection with a selective AgI membrane electrode. The interference of sulfide is totally prevented by its oxidation in the donor line. If applied to potentiometric measurement following gas diffusion separation an intrinsically non-selective metallic silver wire electrode turns out to enable the selective detection of sulfide with high sensitivity and fast response. A new approach for diffusive sampling and on-line detection of gas-phase contaminants is exemplified by the determination of NO_x.     

Flow-injection analysis of serum urea using o-phthalaldehyde and naphthylethylenediamine

An analytical procedure is developed for the determination of urea using flow injection analysis. The methodology is based on the color that develops (lambda(max), 517 nm) when urea reacts with o-phthalaldehyde in the presence of naphthylethylenediamine under acidic conditions. Calibration curves are found to be linear up to 51 mg urea N/dl when the FIA manifold is operated at 42 degrees , utilizing 90-mul samples and a flow-rate of 0.44 ml/min. By manual injection up to 40 samples can be analysed per hour. Recovery yields varied between 95-99%. The within day CV was 0.5-2.2% whereas the day to day CV was 2.1-3.9%. When applied to the analysis of urea in serum samples, excellent correlations (0.998) are obtained when the FIA results are compared with those obtained for the same serum samples analysed by the free urease/Berthelot's and the hospital method (employing the Abbot Bichromatic Analyser). Interferences are observed with sulphur compounds such as sulphanilamide (0.76 mg/dl) as well as with many amino acids but at relatively high concentrations (21 mg/dl).     

HPLC Determination of Glyphosate,Aminomethylphosphonic Acid,and Glufosinate Using a Hypercarb Porous Graphite Adsorbent

A method for the HPLC determination of glyphosate, aminomethylphosphonic acid, and glufosinate using the gradient separation of analytes on a Hypercarb porous graphitized carbon adsorbent and an aqueous solution of ammonium formate and ammonia as a mobile phase is proposed. Analytes are detected using quadrupole and three-quadrupole mass spectrometers. In order to increase the retention of the analytes, the chromatographic column is washed with water before the injection of a sample solution. This procedure results in a three- to fourfold increase in the retention factors of the analytes in comparison with the analogues described in the publications.     

Enzymatic flow-injection determination of urea in blood serum using potentiometric gas sensor with internal nonactin based ISE

Ion-selective electrode with cellulose triacetate membrane containing nonactin is employed for the potentiometric detection of ammonia produced in biocatalytic reaction in flow-injection system with enzyme reactor. The elimination of interferences occurring in the presence of alkali metal ions was achieved by covering a nonactin membrane with outer hydrophobic gas permeable membrane. The obtained flow-injection response to ammonia indicates a possibility of ammonia determination down to 10 microM ammonia. In the flow-injection system for urea determination 200 microl of 10-fold diluted blood serum sample was injected into carrier stream of distilled water merged with TRIS buffer, passed through the urease flow-through reactor and then after merging with NaOH stream delivered to the detector. It was found in several series of natural blood serum samples, that the correction for endogenous ammonia in such a determination is not indispensable.     

Determination of total nitrogen in food by flow injection analysis (FIA) with a potentiometric differential detection system

A flow injection set-up based on potentiometric detection and gas diffusion device for the determination of total nitrogen in food is described. The detection system consisted of two ammonium-sensitive electrodes placed sequentially and each alternately operating as reference electrode. Tubular electrodes without an inner reference solution were prepared with a PVC membrane composed of nonactin in Tris (2-ethylhexyl) phosphate and potassium tetrakis (4-chlorophenyl) borate to reduce the membrane resistance. The food sample digests were inserted into the system, and the ammonium present was converted into ammonia gas. The gas diffused through a gas-permeable membrane to a buffer acceptor stream with a pH that ensured transformation to the ammonium cation, which was potentiometrically detected. Good agreement between FIA results and those provided by the reference procedure was obtained, with relative deviation errors below 5%. Using the proposed system, low reagent consumption is possible, a sampling rate of about 30 samples/h was achieved, as well as a good reproducibility for consecutive injections of the same sample (variation coefficient < 2%). Received: 8 October 1998 / Revised: 7 January 1999 / Accepted: 12 January 1999     

Determination of ammonia in beers by pervaporation flow injection analysis and spectrophotometric detection

A pervaporation flow injection (PFI) method is described for the determination of ammonia in beers. After injecting the sample into a NaOH donor solution, ammonia and other volatiles are transferred in the pervaporation unit from the donor stream to an acceptor stream containing sodium salicylate and nitroprusside, which subsequently mixes with alkaline sodium dichloroisocyanurate to allow the classical Berthelot reaction to take place. The blue-coloured complex formed is monitored spectrophotometrically at 655 nm. A linear calibration curve with a range of 0.1–40 mg l⁻¹ was obtained. The method has a detection limit of 0.05 mg l⁻¹ and is capable of a sampling frequency of 11 h⁻¹ at 4 mg l⁻¹ ammonia. It was applied successfully to the determination of ammonia in synthetic samples and unfiltered lager beers. The advantages of the present method over the ammonia ion-selective electrode method and the PFI system based on mixed indicator detection are discussed.     

Determination of total ammonium-nitrogen and free ammonia in a fermentation medium by sequential injection analysis

Summary Automated methods for the determination of ammonium and ammonia are reviewed, and techniques based on gas diffusion using a semi-permeable membrane were selected for the determination of both total ammonium-nitrogen and free ammonia in fermentation samples. A simple and robust instrument based on sequential injection analysis (SIA) consisting of a piston pump and two selector valves was used. Two different methods of ammonia detection, the Berthelot method and detection using pH-indicators, have been evaluated and compared. The indicator method showed higher reproducibility and the range of determination could easily be adjusted to match the concentrations of the samples. The range of determination could be adjusted from 0.05 mmol/l to 350 mmol/l, depending on choice of acceptor solution. This method has been evaluated with fermentation medium samples and tested on-line in a yeast fermentation process.Dedicated to Professor Dr. Wilhelm Fresenius on the occasion of his 80th birthday     

Determination of titratable acidity and ascorbic acid in fruit juices in continuous-flow systems

Summary Two continuous-flow systems for the determination of titratable acidity and ascrobic acid in fruit juice samples are described. The assemblies permit on-line dialysis of analytes prior to the reaction step, thus improving selectivity and performing sample dilution. Flow systems are built with a channel carrying the donor phase (sample in both determinations) and another channel carrying an acceptor phase, both of them entering the dialyser. The outcoming stream transporting the dialysed sample fills the valve loop, permitting its injection into a carrier stream which continuously passes through the spectrophotometric detector. For the titratable acidity, acceptor phase and carrier are distilled water, the reagent merged with the carrier channel being a buffered solution of bromothymol blue (pH 7). The analytical signal obtained is then monitored at 616 nm. For ascorbic acid, the acceptor phase was a Fe(III) solution, which reacts with the dialysed analyte to form Fe(II). A buffered solution of o-phenanthroline (pH 4.5) is used as carrier, reacting with Fe(II) to give the analytical signal, which is monitored at 510 nm. Chemical and physical parameters are optimized for both systems. The analytical features of the determination are established. Finally, the proposed procedures are compared with the official volumetric AOAC methods for both parameters. The FIA methods turn out to be suitable for a rapid and accurate control of fruit juice samples, compared with the reference methods; additionally they compete advantageously with the volumetric methods in the case of turbid and highly coloured samples.     

Determination of ammonium in Kjeldahl digests by gas-diffusion flow-injection analysis with a bulk acoustic wave-impedance sensor

A novel flow-injection analysis (FIA) system has been developed for the rapid and direct determination of ammonium in Kjeldahl digests. The method is based on diffusion of ammonia across a PTFE gas-permeable membrane from an alkaline (NaOH/EDTA) stream into a stream of diluted boric acid. The trapped ammonium in the acceptor is determined on line by a bulk acoustic wave (BAW)-impedance sensor and the signal is proportional to the ammonium concentration present in the digests. The proposed system exhibits a favorable frequency response to 5.0 x 10(-6)-4.0 x 10(-3) mol l(-1) ammonium with a detection limit of 1.0 x 10(-6) mol l(-1), and the precision was better than 1% (RSD) for 0.025-1.0 mM ammonium at a through-put of 45-50 samples h(-1). Results obtained for nitrogen determination in amino acids and for proteins determination in blood products are in good agreement with those obtained by the conventional distillation/titration method, respectively. The effects of composition of acceptor stream, cell constant of conductivity electrode, sample volume, flow rates and potential interferents on the FIA signals were discussed in detail.     

Optosensing at active surfaces — a new detection principle in flow injection analysis

Reflectance spectrophotometry is applied to flow-injection measurements of pH and the assays of ammonia and urea with the aim of demonstrating the principle and testing the performance of optosensors integrated into microconduits. A novel injection approach, the split-loop technique, is applied. For pH measurements, detection is based on commercial non-bleeding acid-base indicator papers situated in the flow stream at the tip of the fibre optic. Measurements of pH in the range 4–10 are possible at a rate of 120 h^?1. Special attention is given to the physiological pH range; the standard deviation is 0.004 at pH 7.2. For the determinations of ammonia and urea (via urease), a bromothymol blue stream is used with a miniature gas-diffusion device.