Cadmium Reduction Research Articles

Flow-Injection Spectrophotometric Determination of Nitrite and Nitrate in Biological Samples

Nitrite and nitrate in biological samples can act as an indicator of nitric oxide generated in vivo and have attracted a lot of attention as a messenger of diverse physiological processes. For the determination of such nitrogen oxides, a simple and sensitive flow-injection method coupled with a successful pretreatment method was developed. Nitrite and nitrate in sample solution were injected into a carrier solution containing EDTA and ammonium buffer, and flowed into the copperized cadmium (Cd/Cu) reduction column installed on-line. Here the nitrate was reduced to nitrite, and then nitrite was determined spectrophotometrically on the basis of a diazotization/coupling reaction. The detection limits for nitrite and nitrate were 5×10-8 M, and the sampling rate was about 40 samples per hour for nitrate determination. The biological samples, such as serum, plasma and cell culture medium, were deproteinized using a batchwise NaOH-ZnSO4 method before FIA measurement. The NaOH-ZnSO4 deproteinization method was very effective in term of the recovery of nitrate and the maintenance of the reduction efficiency of the Cd/Cu column. The relative standard deviations were 2.1 - 3.1%, and the recoveries were 95 - 100% for the determination of nitrite and nitrate in serum samples through the whole procedure. In the case of cell culture medium, the proposed deproteinization was especially effective for removing the interfering amino acids containing sulfur atom by using the reaction between Zn2+ in deproteinization agent and sulfur atom in analytical substances.

Cerebrospinal Fluid Nitrate Levels in Patients with Multiple Sclerosis

It has been suggested that nitric oxide (NO) could be implicated in the pathogenesis of multiple sclerosis (MS). Recently, two groups reported increased cerebrospinal fluid (CSF) nitrate levels (oxidation product that provides an indirect estimation of NO) in MS patients. However, another group did not confirm these findings. We studied the CSF and plasma levels of nitrate with a kinetic cadmium reduction method in 11 MS patients and 25 matched controls. The CSF nitrate levels and the CSF/plasma nitrate ratio did not differ significantly between the two study groups. Plasma nitrate levels were nearly significantly lower in MS patients. CSF and plasma nitrate levels did not correlate with age at onset and duration of the disease in the patient group. These data suggest that measurement of CSF levels of nitrate is not a marker of the activity of MS.

A unified approach to trace analysis and evaluation of electrode kinetics with fast Fourier transform electrochemical instrumentation

Abstract A microcomputer-based form of Fourier transform instrumentation, which yields potential and frequency-dependent current, admittance and impedance data with excellent signal-to-background ratio over a wide frequency range, is shown to provide state-of-the-art performance in both analytical and kinetic applications of voltammetry. The limit of detection of 5×10−8 to 10−7 M found for the determination of cadmium and lead at a hanging mercury drop electrode, when applying a multi-frequency excitation signal, is equivalent to that reported with single-frequency or time-domain ac, pulse and square-wave polarographs. The highly effective discrimination against the charging current, which is required for trace analysis, can be achieved only by correction of the real part of the admittance data for the uncompensated solution resistance (a parameter readily obtained from impedance analysis of the data). Furthermore, excellent agreement with literature data is achieved when calculating the physico-chemical parameters associated with the cadmium(II) reduction process in different media. The ability to use the same instrument to undertake analytical determinations and characterise the electrode kinetics represents an ideal method for confirming the fidelity of a voltammetric analysis.

FIA evaluation of nitrite and nitrate contents of liver pâtés

A comparative study on the nitrite and nitrate contents of 15 liver pâté brands, in a total of 45 samples (three batches per brand) is presented. The study was conducted on two different kind of pâtés: pork liver pâtés and fowl liver pâtés. For the simultaneous determination of nitrite and nitrate, an automated flow injection system with spectrophotometric detection was used. The manifold was based on the splitting of the flow after injection and subsequent confluence of the flow before reaching the detector, allowing the reduction of nitrate to nitrite in part of the sample plug on an on-line copper cadmium reductor column. Spectrophotometric determination was made after a diazotization coupling reaction. The levels of nitrites and nitrates ranged from 1.07 mg NaNO 2 kg −1 ± 0.43 to 15.9 mg NaNO 2 kg −1 ± 5.2 and from 24.5 mg NaNO 3 kg −1 ± 2.7 to 207 mg NaNO 3 kg −1 ± 14, respectively. The levels of concentration of these constituents were below the allowable limits. A significant dispersion in the results was observed between different brands and within some of the brands.

Spectrophotometric determination of nitrate and nitrite in soil and water samples with a diazotizable aromatic amine and coupling agent using column preconcentration on naphthalene supported with ion-pair of tetradecyldimethylbenzylammonium and iodide

Composite diazotization-coupling reagents containing sulfanilamide (SAM), sulfapyridine (SP) or sulfathiazole (ST) as the diazotizable aromatic amines and sodium 1-naphthol-4-sulfonate (NS) as the coupling agent using column preconcentration on naphthalene-tetradecyldimethylbenzylammonium(TDBA)-iodide adsorbent have been used for the spectrometric determination of trace nitrate and nitrite in soil and water samples. Nitrite ion reacts with SAM in the pH range 2.0–5.0, SP in the pH range 2.0–2.5 and ST in the pH range 2.0–3.3 in HCl medium to form water-soluble colourless diazonium cations. These cations were coupled with NS in the pH range 9.0–12.0 for the SAM system, 9.6–12.0 for the SP system and 8.5–12.0 for the ST system to be retained on naphthalene-TDBA-I material packed in a column. The solid mass is dissolved from the column with 5 ml of dimethylformamide and the absorbance is measured spectrometerically at 543 nm for SAM-NS, 533 nm for SP-NS and 535 nm for ST-NS. Nitrate is reduced to nitrite by a copper-coated cadmium reductor column and the nitrite is then treated with the diazotization-coupling reagent by column preconcentration. The absorbance due to the sum of nitrate and nitrite is measured and nitrate is determined by difference. The calibration graph was linear over the range 2–40 ng NO 2 −-N ml −1 and 1.5–30 ng NO 3 −-N ml −1 in aqueous samples for the SAM and ST systems and 2–48 ng NO 2 −-N ml −1 and 1.5–36 ng NO 3 −-N ml −1 in aqueous samples for the SP system, respectively. The sensitivity, accuracy and precision of the systems decreased in the order ST⪢SAM⪢SP. The detection limits were 1.4 ng NO 2 −-N ml −1 and 1.1 ng NO 3 −-N ml −1 for SAM, 1.6 ng NO 2 −-N ml −1 and 1.2 ng NO 3 −-N ml −1 for SP, and 1.0 ng NO 2 −-N ml −1 and 0.75 ng NO 3 −-N ml −1 for ST, respectively. The preconcentration factors are 8, 5 and 6 for SAM-NS, SP-NS and ST-NS, respectively. Interferences from various foreign ions have been studied and the methods have been applied to the determination of ng ml −1 levels of nitrite and nitrate in soil and water samples. The mean recovery was 95–102% for all three systems.

The Effect of Blocking Additives on the Electrodeposition of Cadmium

Voltammetric and current efficiency studies were made on the effect of benzyl and substituted benzyl alcohols on the competitive deposition of cadmium and evolution of hydrogen. The adsorbed alcohol additives acted as general blocking agents blocking the evolution of hydrogen. In the weakly complexing sulfate system these additives formed complexes with the Cd(II), and in the chloride system they caused a cathodic shift of the cadmium reduction potential, indicating a shift in the distribution of the chloride complex ions to a higher concentration of the more highly complexed species. Electrogenerated hydroxyl ions precipitated cadmium hydroxide in longer chain alcohol films. It was shown that in the presence of these alcohol additives in the chloride system in which a portion of the Cd‐bearing ions are negatively charged, the addition of quaternary ammonium ions, which are known phase‐transfer catalysts, accelerated the rate of reduction of cadmium(II), indicating that ion pairing occurred. On the other hand, these additives either inhibited or did not affect the rate of reduction of cadmium(II) in the sulfate systems in which the cadmium ions are present as positively or neutrally charged species. The fact that the ion pairing and acceleration occurs with the negative chloride complex, even though this complex is present in very small concentrations in the bulk of the solution, indicates that the presence of the low‐dielectric‐constant alcohol at the electrode interface has increased the concentration of these negative complexes in the neighborhood of the electrode.

Nitro-L-arginine inteferes with the cadmium reduction of nitrate/griess reaction method of measuring nitric oxide production.

Nitro-L-arginine is used as an inhibitor of nitric oxide synthase in many biological Systems. Nitric oxide (NO) is unstable and degrades to nitrite NO(2)- and nitrate NO(3)-. The colorimetric reaction of N0(2)- with Griess reagent is commonly used to measure NO(2)-. NO(3)- may be measured as NO(2)- following reduction by cadmium or cadmium/copper. We found that bradykinin increased the formation of NO(2)- by bovine coronary endothelial cells. Nitro-L-arginine further increased the formation of NO(2)-. This increase is due to the interference of nitro-L-arginine in determination of NO(3)- by the cadmium reduction to NO(2)- and Griess reagent reaction. Incubation of nitro-L-arginine with cadmium or cadmium/copper produced a product that reacts with Griess reagent to form a compound that has an absorption spectrum identical to the product formed by NO(2)- and Griess reagent. Caution must be exercised when using the NO(2)-/NO(3)- measurement by the Griess reaction to assess inhibition of nitric oxide synthase by nitro-L-arginine.

Oxidative detection of nitrite at an electrocatalytic [Ru(bipy) 2poly-(4-vinylpyridine) 10Cl]Cl electrochemical sensor applied for the flow injection determination of nitrate using a Cu/Cd reductor column

An electrochemical sensor developed from a redox polymer [Ru(bipy) 2(PVP) 10Cl]Cl (where bipy is 2,2′-bipyridyl and PVP is poly-(4-vinylpyridine)) modified electrode for nitrite detection is described. The modified electrode acts as an efficient electrocatalyst for nitrite oxidation with amplification of electrode current by at least a factor of 3 compared with unmodified electrodes. The electrocatalytic reaction occurs throughout the polymer film in a three dimensional reaction zone and may be classified as an L k mechanism. The second order rate constant for the cross exchange reaction is 3.02 (± 0.20) × 10 1 mol −1 dm −3 s −1. The sensor is applied to the determination of nitrate in a flow injection apparatus following the heterogeneous reduction of this species to nitrite at a copperised cadmium reductor column. The sensor exhibits a linear response range extending over three orders of magnitude from 1.0 × 10 −5 to 1.0 × 10 −2 mol dm −3 nitrate with a limit of detection (signal-to-noise = 2) of 5.0 × 10 −6 mol dm −3. The coefficient of variation for measurement of 1.0 × 10 −3 mol dm −1 nitrite is 0.42% ( n = 20). No passivation of the electrode surface was evident after over 200 repeat determinations. The system was applied to the determination of nitrate in commercial fertiliser compound.

Simultaneous assay of nitrite, nitrate and chloride in meat products by flow injection.

A flow injection (FI) analytical method for the simultaneous assay of nitrite, nitrate and chloride in meat products is reported. The method is based on the potentiometric determination of chloride using a tubular ISE and on the spectrophotometric determination of nitrite. The FI system consisted in splitting the flow after potentiometric detection using a tubular detector and the subsequent confluence of the flow before reaching the spectrophotometric detector. This allowed the reduction of nitrate to nitrite in part of the sample plug on an on-line copper cadmium reductor column. Since each channel had a different residence time, two peaks were obtained for nitrite and nitrite plus nitrate. Spectrophotometric determination was made after a diazotization-coupling reaction. The results obtained were in good agreement with reference procedures and showed adequate precision (RSDs less than 6% for chloride and nitrite and 2% for nitrate). A high sampling rate was obtained (120 determinations per hour corresponding to 40 samples per hour).

Use of reflectometry for determination of nitrate‐nitrogen in well water

Abstract Recurrent monitoring of water wells is necessary to ensure that nitrate‐nitrogen (NO3‐N) concentrations in groundwater do not exceed 10 mg/L, the maximum contaminant level set by the U.S. Environmental Protection Agency. Continuous chemical analysis is often a time consuming and expensive process. A recently developed ‘Reflectoquant Analysis System’, which employs reflectometry techniques, may offer a simple and accurate method for NO3‐N analysis. The objective of this study was to evaluate the ‘Reflectoquant Analysis System’ as an alternative method for determination of NO3‐N in well water. Water samples were collected from 42 wells in Oklahoma. The samples were analyzed using the ‘Reflectoquant Analysis System’, automated cadmium reduction (Griess‐Ilosvay), ion chromatography, and phenoldisulfonic acid procedures. The linear range of the ‘Reflectoquant Analysis System’ is 1.1 to 50.6 mg/L NO3‐N. Samples exceeding this range must be diluted before analysis is performed. Excluding two wells where...

Simultaneous Determination of Nitrate and Nitrite in Biological Samples by Multichannel Flow Injection Analysis

An automated method for the simultaneous determination of nitrite and nitrate in biological samples by using a multichannel flow injection analyzer has been developed. The method was based on the reaction of nitrite with Greiss reagent. The sample solution was injected and equally divided into two channels; channel one (1) represented total nitrite obtained by cadmium reduction of nitrate to nitrite while channel two (2) represented only nitrite. The absorbance of the color product was measured by photometric detectors with 540-nm filters. This method combines high reproducibility of sample introduction via flow injection and sensitivity of spectrophotometric detection. The detection limit is 25 nMfor both nitrite and nitrate. The chemistry manifolds are constructed of Teflon tubing which, along with a low-pressure Flowfit connector system, provides for low maintenance, ease of use, and high sample throughput. We demonstrated that the system can be used for the determination of both nitrate and nitrite in a variety of biological samples as well as a comparison of the results from this system and the HPLC system.

Bray and Kurtz phosphorus (p1), 1m potassium chloride, and saturated calcium oxide extraction of soil nitrate comparison to 2m potassium chloride

Abstract Labor and laboratory expense could be reduce if soil extractable nitrate (NO3) was extracted simultaneously with other routinely used soil extractants. This study was conducted to compare 1M potassium chloride (KCl), saturated calcium oxide (CaO), and Bray and Kurtz P1‐extractable soil NO3, the proposed new method to 2M KCl. Nitrate was extracted from 140 selected soils with 1M KCl, saturated CaO, Bray and Kurtz P1, and 2M KCl extractants and analysed by cadmium reduction with a flow injection analyser. Nitrate extracted with 2M KCl was used as the standard. When 1M KCl, saturated CaO, and Bray and Kurtz P1 was regressed against 2M KCl, the slopes were equal to 1.0 and the intercepts were equal to 0.0 with r2>0.99. No detectable differences in extractable NO3 were measured between 1M KCl, saturated CaO, Bray and Kurtz Pl, and 2M KCl extraction procedures. Bray and Kurtz P1 can be used to simultaneously extract phosphorus (P) and NO3.

Sample Pretreatment with Nitrate Reductase and Glucose-6-Phosphate Dehydrogenase Quantitatively Reduces Nitrate While Avoiding Interference by NADP+ When the Griess Reaction Is Used to Assay for Nitrite

An assay for the simultaneous measurement of nitrite and nitrate, products of nitric oxide metabolism, is described. Others have reported pretreating sample by using nitrate reductase (NR) and NADPH to reduce endogenous NO−3 before assaying the resultant NO−2 using the Griess reaction. However, we found that the NADP+ formed during pretreatment interfered with the Griess reaction when NADPH was used at concentrations necessary to drive the NR reaction. For instance, 500 μM NADP+ in 100 μM NaNO−3 (without NR) causes a 90% interference with the formation of Griess reaction product. To limit interference, we modified the method by decreasing the NADPH concentration to 1 μM. NADPH was regenerated by coupling the NR reaction with that catalyzed by glucose-6-phosphate dehydrogenase (GD). Using this method, NaNO−3 standard curves were linear up to 100 μM and coincided with control curves obtained using NaNO−2 incubated in parallel. Addition of urine up to a strength of 20% did not interfere with the assay. Comparison with an alternative assay based on cadmium reduction resulted in the following linear regression: [Cd method] = 0.915∗[NR-GD method] + 0.37, r2 = 0.997. Coupling GD to NR to recycle NADPH allows this cofactor to be used at a low concentration so that interference with the Griess reaction is negligible.

Endotoxin-induced nitric oxide production in pulmonary artery endothelial cells is regulated by cytokines.

L-Arginine is the sole precursor of nitric oxide (NO). Bacterial lipopolysaccharide (endotoxin) (LPS) stimulates carrier-mediated L-arginine transport in porcine pulmonary artery endothelial cells (PAECs) through an autocrine pathway that involves interleukin-1 alpha (IL-1 alpha) and tumor necrosis factor alpha (TNF-alpha). To determine if Escherichia coli LPS stimulates NO synthesis in PAECs and, if so, if LPS stimulation of NO production is also mediated by autocrine secretion of IL-1 alpha and TNF-alpha. Monolayers of PAECs were incubated with various concentrations of LPS, recombinant human TNF-alpha, or IL-1 alpha, and total nitrate-nitrite accumulation was measured at different time points with the Greiss reagent following cadmium reduction. Release of TNF-alpha and IL-1 alpha release by LPS-stimulated PAECs were measured using the WEHI (for TNF-alpha) and A375.S2 (for IL-1 alpha) bioassays. The PAECs were then incubated with saline solution or LPS in the presence or absence of either a polyclonal antibody to human TNF or IL-1 receptor antagonist, and nitrate-nitrite accumulation was measured at 48 hours. Production of NO by PAECs was increased 230% by LPS (1 microgram/mL), 350% by TNF-alpha (1000 U/mL), and 240% by IL-1 alpha (1000 U/mL) (P < .05 vs control). The LPS-stimulated NO production was inhibited by IL-1 receptor antagonist (100 micrograms/mL) or antibody to TNF (10 micrograms/mL) to control levels (P < .05 vs LPS; difference vs saline solution was not significant). The LPS-stimulated TNF-alpha secretion by PAECs and TNF-alpha activity were maximal at 6 hours (400 +/- 42 pg/mL). The IL-1 alpha activity was not detectable in LPS-stimulated PAECs by the A375.S2 bioassay. Endotoxin, TNF-alpha, and IL-1 alpha stimulated NO synthesis in PAECs. Endotoxin-stimulated NO synthesis through an autocrine pathway involving the cytokines TNF-alpha and IL-1 alpha.

Determination of Nitrate in the Presence of Nitrite in Natural Waters by Flow Injection Analysis with a Non-Quantitative On-Line Cadmium Reductor

Abstract Several analytical problems in the determination of nitrate using flow injection analysis (FIA) coupled with an on-line Cd reductor have been studied. It was found difficult to prepare a nearly 100%-efficient copperized Cd reductor which maintains its efficiency over a lengthy period. Instead, the use of a narrow and lower efficiency Cd coil is recommended because it is more stable and therefore more suitable for FIA. Since the conversion of nitrate to nitrite is not quantitative, results for nitrate tend to be over-estimates when nitrite is also present. This problem has been solved by using a simple correction scheme to compensate for the effect of nitrite, thus enabling the correct nitrate concentration to be evaluated. The validity of the correction procedure has been confirmed by running a series of known standards containing both nitrate and nitrite with three types of FIA manifolds. Results for nitrate were accurate for fresh and saline waters even when the co-existing nitrite concentratio...

Analysis of total dissolved nitrogen in natural waters by on-line photooxidation and flow injection

A sensitive automated method for the determination of total dissolved nitrogen (TDN) is described. This method is based on the complete conversion of inorganic and organic nitrogen species to nitrate by photooxidation in an alkaline peroxodisulphate medium. Following on-line photooxidation, sodium metabisulphite is added to reduce excess peroxodisulphate reagent, and a portion of the resultant solution is injected into a flow-injection manifold for the measurement of nitrate by cadmium reduction to nitrite, diazotization, coupling and detection at 540 nm. The method is comparatively fast (ca. 25 determinations h −1, has a detection limit of 0.03 mg 1 −1 N, and is linear up to 3 mg 1 −1 N. The PTFE tubing photoreactor is simple to construct, and because detection is by visible spectrophotometry, the method could be readily employed using either commercial or low-cost laboratory-built flow-injection analysis systems.

Determination of total nitrogen in water samples by means of high-pressure bombs and ion chromatography

The standard method for the determination of organic nitrogen in water samples in the past has been based on the Kjeldahl digestion technique that converts organic nitrogen into ammonia, which can then be determined along with any ammonia originally present. However, this method is time consuming and the accuracy is variable because the effect of interferences causes unreliable results, especially in complex matrices. An alternative method for the determination of organic nitrogen is the alkaline peroxodisulphate digestion technique. This oxidizes all nitrogen in the sample using potassium peroxodisulphate in a strongly alkaline environment under high pressure and temperature. Nitrate is the sole product and can easily be determined by ion chromatography or by other methods ( e.g., cadmium reduction method). A method for total nitrogen determination was developed using 23-ml high-pressure bombs with potassium peroxodisulphate and sodium hydroxide to oxidize the organic nitrogen to nitrate. Urea and ammonium chloride were used as nitrogen compounds for calibration. The method was checked with the Kjeldahl method, showing good agreement (R.S.D. = 4.62%). Studies of digestion time were carried out to determine the optimum time in the pressure vessel. The results were checked with the cadmium reduction method (R.S.D. = 3.62%) for natural water samples. The recoveries with urea and ammonium chloride reagents were higher than 90%.

Spectrophotometric Determination of Nitrate in Baby Foods: Collaborative Study

A collaborative study was conducted of a spectrophotometric method for determination of nitrate after cadmium reduction to nitrite in baby foods containing meat. Thirty-one municipal and 2 industrial food laboratories participated in the study. The study design involved 2 baby food matrices. Samples of both matrices were prepared at 3 concentration levels between 52 and 309 mg NaNO3/kg as blind duplicates. A blank without added nitrate was also included. The outlier percentage of the results was very low (4.3%). It was typical for the method that recoveries were slightly > 100%. Recoveries for baby foods varied between 113.3 and 116.9%, and were acceptable for control purposes. The relative standard deviations for repeatability were 5.0-18.1%. The relative standard deviations for reproducibility were 8.3-21.6%. Three collaborators also evaluated liquid chromatographic technique for nitrate determinations. These preliminary results are presented but are not analyzed statistically. The spectrophotometric method was adopted first action by AOAC INTERNATIONAL.

Indirect Flow Injection Spectrophotometric Determination of Molybdenum(VI) Using an On-Line Cadmium Reductor

An indirect flow injection spectrophotometric method is described for determination of molybdate(VI). The method is based on reducing Mo(VI) in an on-line copperized cadmium reductor and then oxidizing it by reaction with Fe(III). The resulting Fe(II) reacts with ferrozine to form a complex whose absorbance is measured at 562 nm. The linear response range and detection limit (3× the standard deviation of blank) are 1.0-9.4 × 10−5 and 1.1 × 10−6M. respectively. The interference effects of some other ions and their removal are studied. The cadmium reductor can be used for more than 140 injections without any change in response.

Mass transfer and kinetic behaviour of chelated cadmium solutions

The effect of the chelating agents nitrilotriacetic acid (NTA) and ethylenediaminetetraacetic acid (EDTA) on the electrodeposition of cadmium from solutions was studied. Experimental results show that the presence of a chelating agent severely inhibits cadmium reduction. The mass transfer and kinetic behaviour of cadmium ions in chelated solutions (NTA and EDTA) were investigated. The observed overall diffusion coefficients of the chelated cadmium solutions, using the simple Levich equation, were found to be a function of the solution pH and the chelating agent. The relevant kinetic properties, such as the reaction order, α z, k f 0 and the equilibrium potential, were measured for cadmium ions in the chelated solutions. The diffusion coefficients of the more stable chelated cadmium species were found to be less than those of the less stable chelated cadmium species, thus causing more interference in the diffusion step.