University of North Florida
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Stuart Chalk, Ph.D.
Department of Chemistry
University of North Florida
Phone: 1-904-620-1938
Fax: 1-904-620-3535
Website: @unf

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Classification: Biological fluid -> saliva

Citations 17

"Determination Of Mercury In Saliva With A Flow Injection System"
Anal. Chim. Acta 1996 Volume 320, Issue 2-3 Pages 171-176
T. Guo*, J. Baasner, M. Gradl and A. Kistner

Abstract: The method involved treating samples with a brominating agent followed by online addition of KMnO4 to convert organic Hg to inorganic Hg ions. Hg was determined by flow injection cold vapor AAS. A Perkin-Elmer FIMS-400 system (schematic given) with two pumps and tubing of 1.52 mm i.d. was used. The prefill time was 15 s, and the sampling time 10 s. Saliva (0.5 ml) was mixed with 0.5 mL 32% HCl and 0.2 mL bromate/bromide reagent (2 g KBr/0.56 g KBrO3 in 25 mL H2O) and diluted to 5 mL. Portions were injected into the 3% HCl carrier and mixed with 0.25% KMnO4 and 0.2% NaBH4 streams and the products were carried in Ar to the detector. Results were compared with those obtained by a batch method. The detection limit was 0.05 µg/l Hg in saliva diluted 1:10 and the calibration graph was linear for up to 20 µg/l; the dynamic range was up to 40 µg/l. The RSD (n = 11) at 1 µg/l was 2%. Sample throughput was 80 measurements per hour.
Mercury Spectrophotometry

"Determination Of Nitrite Based On Mediated Oxidation At A Carbon Paste Electrode Modified With A Ruthenium Polymer"
Talanta 1992 Volume 39, Issue 4 Pages 443-447
Thomas J. O'Shea, Dónal Leech, Malcolm R. Smyth and Johannes G. Vos

Abstract: Carbon paste electrodes were modified with [Ru(bpy)2(PVP)10Cl]Cl [I; where bpy = 2,2'-bipyridyl and PVP = poly(4-vinylpyridine)] and used for the amperometric detection of NO2-. The surface modifier lowered the overpotential for NO2- oxidation and thus allowed its determination at a lower potential. A 5% I loading and a monitoring potential of 0.8 V gave rectilinear calibration graphs from 50 nM to 0.5 mM NO2- and a detection limit of 30 nM-NO2-. The between-electrode precision was 11.4% (n = 10) and the response was relatively stable over 5 days, with a signal diminution of 8%. In flow injection studies with 0.1 M NaNO3 as supporting electrolyte (1 mL min-1), the rectilinear range was 0.2 µM to 1 mM with a detection limit of 80 nM. In the determination of 0.2 mM NO2- over 90 min (n = 100), the coefficient of variation was 2.9%. The electrode was used to determine NO2- in saliva by flow injection analysis.
Nitrite Electrode Electrode Amperometry

"Amperometric Detection With Microelectrodes In Flow Injection Analysis: Theoretical Aspects And Application In The Determination Of Nitrite In Saliva"
Talanta 1998 Volume 47, Issue 3 Pages 651-658
Vânia Mori and Mauro Bertotti*

Abstract: The construction of a wall-jet cell with amperometric detection using a set of disc electrodes whose radii ranged from 5 to 750 gm has been proposed. The influence of some experimental parameters like flow rate and electrode radius on hydrodynamic voltammograms recorded for a 0.5 mmol L-1 potassium ferrocyanide solution also containing 0.1 mol L-1 KCl has been discussed. Some considerations regarding the current signals obtained from flow injection experiments using both a 5- and a 750-µm radius platinum electrode were carried out in order to achieve the lowest limit of detection, a value of 0.03 µmol L-1 ferrocyanide being calculated by using the 5-µm radius microelectrode as amperometric detector. The wall-jet cell has been used in the determination of nitrite in saliva by quantifying the triiodide formed in the reaction of the analyte with excess iodide in acidic medium. A 12.5-µm platinum disk microelectrode maintained at + 0.2 V vs. Ag/AgCl was used as amperometric detector. Peaks obtained in diagrams after injection of diluted saliva to the carrier stream containing 0.1 mol L-1 sulfuric acid and 20 mmol L-1 potassium iodide were compared to an anal. curve obtained in the same conditions (τ2 = 0.997) for a nitrite concentration. in the range 1-10 µmol L-1. The concentration. of nitrite in the saliva sample after the appropriate correction for dilution was found to be 2.3 ppm (0.05 mmol L-1), in a good agreement with results obtained by using a standard spectrophotometric procedure (2.5 ppm). The limit of detection of the method was calculated as 0.2 µmol L-1, and the reproducibility was checked by measuring the peak current for 19 injections of 10 µM nitrite, the standard deviation being 3.7%.
Nitrite Amperometry Electrode Electrode Indirect Method comparison Apparatus

"High Performance Liquid Chromatography Using A Color-forming Agent As A Component Of The Mobile Phase. Separation And Determination Of Magnesium And Calcium In Clinical Samples With O-cresolphthalein Complexan"
Analyst 1988 Volume 113, Issue 2 Pages 247-250
Jun'ichi Toei

Abstract: The cited determination was carried out on a column (5 cm x 4 mm) of TSK-GEL SP-2 SW. The optimum mobile phase (1 mL min-1) contained 0.2 M KCl, 0.02 M acetic acid - Na acetate buffer (pH 4.0) and 0.1 mM o-cresolphthalein complexan. Detection was at 575 nm after mixing the column eluate with 0.4 M NH3 - NH4Cl buffer (pH 10.4). Pre-treatment of samples (milk, urine, serum or saliva) removed protein-bound Ca; pre-treatment on Toyo-Pack SP cartridges to remove free Ca was unsuccessful. Transition metals and Na did not interfere at physiological concentration. The coefficient of variation (n = 7) for 2.1 ppm of Mg and 28.9 ppm of Ca in milk were 1.41 and 1.26%, respectively.
Calcium Magnesium Clinical analysis Spectrophotometry Interferences Post-column derivatization

"Flow Injection Amperometric Determination Of Thiocyanate And Selenocyanate At A Cobalt Phthalocyanine Modified Carbon Paste Electrode"
Analyst 1994 Volume 119, Issue 7 Pages 1607-1612
Efstathios G. Cookeas and Constantinos E. Efstathiou

Abstract: Cyclic voltammetry was performed with a Tacusel Model PRG-5 polarograph. The three-electrode cell comprised a C paste electrode (CPE) or a cobalt phthalocyanine CPE electrode, a SCE reference electrode and a Pt-wire auxiliary electrode. Supporting electrolyte was Britton-Robinson buffer solution, 0.04 M in each component). The flow injection system comprised a Tecator FIAstar Model 5020 analyzer. coupled to a lab-built Plexiglas wall-jet type amperometric flow detector (schematic shown). The linear response and detection limit were 2-50 µM- and 0.6 µM-thiocyanate, respectively, with an RSD of 2-3% for 3-50 µM-thiocyanate. For selenocyanate, the corresponding figures were 0.2-20 µM and 0.1 µM with an RSD of 2-3% for 0.5-20 µM. Results obtained on applying the method to saliva samples agreed well with those obtained by the standard spectrophotometric method and literature data.
Thiocyanate ion Selenocyanate Voltammetry Electrode Spectrophotometry Amperometry Tecator

"Electrochemical Sensor Of Nitrite Based On An Inorganic Film Modified Glassy Carbon Electrode"
Microchim. Acta 1993 Volume 111, Issue 1-3 Pages 63-70
Zhiqiang Gao, Mingxian Zi, Yimin Zhang, Guangqiang Wang and Zaofan Zhao

Abstract: A thin-film of copper heptacyanonitrosylferrate was electrodeposited on a vitreous carbon electrode which was then used to study the electrocatalysis of nitrite in aqueous solution Cyclic voltammetry of the modified electrode in nitrite solution showed that both oxidation and reduction were catalyzed and that the electrocatalytic currents were controlled by the diffusion of nitrite. When applied as an amperometric sensor in a FIA system, the modified electrode permitted detection at -0.55 V, more than 500 mV lower than at the naked electrode surface. Under these conditions the calibration graph was rectilinear for 1 µM to 1 mM nitrite and the detection limit was 0.3 µ-scm. The coefficient of variation (n = 50) in the determination of 50 µM-nitrite was 3.9%. Common anions such as Cl-, SO42-, NO3- and PO43- did not interfere but H2O2 and dissolved O interfered seriously; ascorbic acid may also interfere. The modified electrode was used for several days without substantial losses in sensitivity and reproducibility. The procedure was applied to the determination of nitrite in saliva and nitrate.
Nitrite Amperometry Electrode Electrode Sensor Interferences

"Manganese(III) - Porphyrin-based Thiocyanate-selective Membrane Electrodes: Characterization And Application In Flow Injection Determination Of Thiocyanate In Saliva"
Electroanalysis 1989 Volume 1, Issue 6 Pages 477-484
D. V. Brown, N. A. Chaniotakis, I. H. Lee, S. C. Ma, S. B. Park, M. E. Meyerhoff*, R. J. Nick, J. T. Groves

Abstract: The electrode membrane comprised 1% of 5,10,15,20-tetrakis-(2,4,6-triphenyl)porphyrinatomanganese(III) chloride, 66% of dibutylsebacate and 33% of PVC; the internal buffer solution contained 50 mM 2-morpholinoethanesulfonic acid - 10 mM NaCl (adjusted to pH 5.5 with 1 M NaOH), and also contained 50 µM NaSCN. The electrode responds preferentially to SCN- over inorganic (e.g., ClO4-, Cl- and Br-) and organic (e.g., ascorbate, citrate and urate) ions often present in biological samples. Whereas the selectivity of the electrode for SCN- was different in a non-equilibrium flow system compared with that under equilibrium conditions, its response allowed determination of SCN- in saliva by flow injection analysis. The carrier stream was the cited buffer (without addition of NaSCN). From 0.01 to 1 mM SCN- could be determined. Results showed good agreement with those of a colorimetric method.
Thiocyanate ion Electrode Electrode Method comparison

"Liquid Chromatographic Analysis Of Carboxylic Acids Using N-(4-aminobutyl)-N-ethylisoluminol As Chemiluminescent Label: Determination Of Ibuprofen In Saliva"
J. Chromatogr. B 1993 Volume 615, Issue 1 Pages 97-110
O. M. Steijger, H. Lingeman and U. A. Th. Brinkman, J. J. M. Holthuis, A. K. Smilde and D. A. Doornbos

Abstract: N-(4-Aminobutyl)-N-ethylisoluminol was used for labelling of carboxylic acids. The derivatization reaction was carried out with 1-hydroxybenzotriazole as pre-activator of the carboxylic acid function and N-ethyl-N'-(3-dimethylaminopropyl)carbodiimide as the coupling reagent. Optimum conditions for the derivatization were determined by using factorial design analysis, with ibuprofen as the test compound. Chemiluminescence detection was carried out using a post-column online electrochemical hydrogen peroxide generation system and the addition of microperoxidase as the catalyst. The detection limit of derivatized ibuprofen in human saliva was 0.7 ng per 0.5 mL of saliva, with a recovery of 96.1±1.3%. The method was linear over at least three decades (2.5 ng to 2.5 µg) and the repeatability was satisfactory (RSD = 5.2% at the 25 ng level; n = 4).
Carboxylic acids Ibuprofen LC Chemiluminescence Factorial design

"Determination Of BAY Y 3118, A Novel 4-quinolone, In Biological Fluids Using High Performance Liquid Chromatography And Photothermal Post-column Derivatization"
J. Chromatogr. B 1993 Volume 616, Issue 1 Pages 87-93
Hans Guenter Schaefer

Abstract: Plasma was diluted with 0.05 M H3PO4 and centrifuged; saliva and urine were diluted with 0.1 M phosphate buffer of pH 7.5. Portions of the resulting solution were injected on to a column (25 cm x 4.6 mm) of Nucleosil 100 C18 (5 µm) operated at 50°C and protected by a similarly packed guard column with acetonitrile/[0.1 M tetrabutylammonium bromide/0.05 M H3PO4 (1:1)] (19:81; pH 2) as mobile phase (1 ml/min). Post-column derivatization of the eluate was performed, utilizing either photolysis at 254 nm in a Beam Boost reaction unit or combined thermolysis and photolysis in a laboratory-made reactor (details given). Fluorimetric detection was performed, employing excitation at 277 nm and an emission cut-off filter (418 nm). Calibration graphs were rectilinear from 0.01-2.0 mg/l of BAY y 3118 (I) in plasma and 1-500 mg/l of I in urine. Intra- and inter-day RSD were from 3.3-5.1% and 2.6-9.8%, respectively, for plasma and from 0.5-3.8% and 2.5-6.9%, respectively, for urine. The method was used to determine the pharmacokinetic behavior of I in a healthy human volunteer. A reversed-phase high performance liquid chromatographic (HPLC) method that allows the sensitive and selective quantification of a novel 4-quinolone (BAY y 3118, I) in biological fluids is described. After sample dilution with 0.05 M phosphoric acid (plasma) or 0.1 M phosphate buffer pH 7.5 (urine), samples can be directly injected into the HPLC system. Prior to fluorescence detection, I is decomposed to fluorescence compound(s) by post-column derivatization utilizing either photolysis (Beam Boost reaction unit) or a combination of thermolysis and photolysis (laboratory-made post-column reactor). Compared with fluorescence detection alone, derivatization increases the signal intensity (about 80-fold) and the selectivity of the detection significantly. Concentrations down to 0.01 mg/l could be quantified in biological fluids. Only thermolysis was not able to decompose I to fluorescence products. Investigations on the stability of I in plasma and urine demonstrate good stability under the different conditions tested. The method was applied to human plasma and urine samples from a subject after a single oral dose of 100 mg of I.
4-Quinolone HPLC Fluorescence Post-column derivatization

"Rapid And Sensitive Determination Of Nitrite In Foods And Biological Materials By Flow Injection Or High Performance Liquid Chromatography With Chemiluminescence Detection"
J. Chromatogr. A 1994 Volume 673, Issue 1 Pages 77-84
Nrisinha P. Sen*, Philander A. Baddoo and Stephen W. Seaman

Abstract: Cured meats and baby foods were homogenized, extracted with water, and the extract processed and filtered as described by Sen and Donaldson (cf. J. Assoc. Off. Anal. Chem., 1978, 61, 1389). Urine and saliva were analyzed directly. All samples were diluted (2-100 fold) with HPLC mobile phase. In the HPLC method, the sample (50-100 µL) was injected into a mobile phase (1 or 2 ml/min) of 0.05 M KH2PO4 of pH 6/5 mM tetrabutylammonium hydrogensulfate and passed through a column (25 cm x 4.6 mm i.d.) of Techsil C18 (5 µm). The nitrite-containing eluate was mixed with 10% H2SO4 in acetic acid and then with 10% KI (0.5 ml/min) and the solution was transported to a coiled PTFE tubing reactor kept at 70°C. Chemiluminescence detection of the NO thus formed was effected with a thermal energy analyzer.. The flow injection method was performed in the same way, but without the HPLC column. A diagram of the system used is given. The calibration graph was linear from 1-1000 ng of nitrite; the detection limit was 0.1 ng. The RSD (n = 4 or 5) was 0.2-4%. Recoveries were 92-108%.
Nitrite HPLC Chemiluminescence

"Determination Of Oxidizable Inorganic Anions By High Performance Liquid Chromatography With Fluorescence Detection And Application To The Determination Of Salivary Nitrite And Thiocyanate And Serum Thiocyanate"
J. Chromatogr. B 1988 Volume 424, Issue 1 Pages 29-37
Shinzo Tanabe, Michie Kitahara, Masashi Nawata and Kouji Kawanabe

Abstract: Sulfide, SCN-, S2O32- and NO2- were determined by HPLC on a column (15 cm x 4 mm) of TSK gel QAE-2SW (5 µm) with a mobile phase (0.8 mL min-1) of 0.05 M NaNO3 and post-column derivatization, at 80°C for 15 s, with 0.75 M H2SO4 containing 0.25 mM Ce(SO4)2 and 0.003% of sodium bismuthate. Fluorescence detection was at 360 nm (excitation at 260 nm). The limits of detection were 0.1, 0.1, 0.3 and 0.8 nmol for NO2-, SCN-, S2O32- and S2-, respectively. The calibration graphs were rectilinear for 10 nmol per 10 µL injected and the coefficient of variation was <8%. The method was applied in the determination of NO2- and SCN- in saliva and SCN- in serum. Recoveries of NO2- and SCN- were 99.3 and 99.1%, respectively, from saliva and recovery of SCN- from serum was 63%.
Anions, inorganic Nitrite Sulfide Sulfite Thiocyanide HPLC Fluorescence Heated reaction Post-column derivatization

"Enzymatic Determination Of Ethanol In Saliva By Flow Injection Analysis"
J. Pharm. Biomed. Anal. 1987 Volume 5, Issue 7 Pages 701-706
P. Linares, J. Ruz, M. D. Luque De Castro and M. Valcarcel

Abstract: The method described is based on the oxidation of ethanol(I) by NAD+ in the presence of alcohol dehydrogenase. Saliva (50 µL) was diluted to 5 mL with K4P2O7 buffer solution (pH 9.0). Sample solution and enzyme - NAD+ solution were injected simultaneously into two buffer streams; the streams were merged and stopped in the flow cell and the reaction was monitored at 340 nm or fluorimetrically at 460 nm (excitation at 340 nm). A Tecator flow injection analysis 5020 analyzer. was used. Calibration graphs were rectilinear from 2.5 to 15 and 1 to 20 µg mL-1 with spectrophotometric and fluorimetric detection, respectively, with corresponding determination limits of 0.47 and 0.26 µg mL-1; the coefficient of variation were 0.68 and 0.5%. The sampling rate was 40 h-1.
Ethanol Fluorescence Spectrophotometry Immobilized enzyme Merging zones Stopped-flow Tecator

"Fluorimetric Flow Injection Analysis Of Saliva For Nitrite"
Anal. Sci. 1987 Volume 3, Issue 6 Pages 549-551

Abstract: Saliva (100 to 150 mg) is diluted to 20 mL with water, ultrasonically agitated and centrifuged. A portion of the supernatant solution (210 µL) is injected into a carrier stream of water (0.6 mL min-1), which is successively mixed with 6 mM 4-hydroxycoumarin in acetonitrile - 4 M HCl (1:1), 40 mM Na2S2O3 and 3.5 M NaOH at 0°C, room temperature and 100°C, respectively, to give 3-amino-4-hydroxycoumarin. Fluorescence is measured at 455 nm (excitation at 345 nm). Recovery of added NO2- is in the range 95 to 102%. The method is calibrated up to 10 µg mL-1, with a detection limit of ~2 ng mL-1 and coefficient of variation (n = 8) of 2.8 and 1.7% at 20 and 100 ng mL-1, respectively. The sampling rate is 30 h-1.
Nitrite Fluorescence Heated reaction

"Automated Continuous-flow Radio-immunoassay For Salivary Oestriol"
Clin. Chem. 1984 Volume 30, Issue 10 Pages 1678-1682
PL Truran and GF Read

Abstract: The assay is based on a manual method described previously (cf. Anal. Abstr., 1983, 44, 6D157) which involves use of Sepharose-coupled antiserum and 125I-labelled radio-ligand. The assay is automated by using a continuous-flow system ('Southmead System'; cf. Ismail et al., Ibid., 1982, 42, 3D171); bound and free radio-ligand fractions are separated by online filtration. Performance of the assay was reliable over 6 months of regular use, and compared favourably with that of the manual assay. The coefficient of variation were 7.5 to 9.6%. Drift (0.5 nM per 100 samples) and sample carry-over (3%) were acceptable and the sample throughput was 55 h-1.
Oestriol Clinical analysis Radiochemical Radioimmunoassay

"Coupled Reactions For The Determination Of Analytes And Enzymes Based On The Use Of Luminescence"
J. Biolumin. Chemilumin. 1989 Volume 4, Issue 1 Pages 423-435
A. Roda, S. Girotti, S. Ghini, G. Carrea

Abstract: The use of immobilized luminescent enzymes, viz, Photinus-luciferin 4-monooxygenase (ATP hydrolysing) (I), alkanal monooxygenase (FMN-linked) (II) and peroxidase coupled with analyte specific enzymes for analysis of serum and saliva is discussed. Enzymes were immobilized on nylon tubes as described by Hornby and Goldstein (Methods in Enzymology, 1976, 44, 118) or on methacrylate beads. For determinations involving a coupled enzyme reaction with use of II, the analyte-specific enzyme and II were immobilized on separate nylon tubes. For determination of lactate dehydrogenase (III) by this method, the calibration graph was rectilinear from 1 to 500 iu L-1 of III at 37°C and 3 to 2000 iu L-1 at 27°C. Recoveries were 92 to 110% and the coefficient of variation were 10%. Results compared well with those by spectrophotometry. The enzymes used, working ranges, detection limits and precision for 17 analytes are tabulated. ATP and ADP were determined in the pmol range with used of I immobilized on 0.5 to 1 m of nylon tubing.
Luminescence Enzyme Nylon Immobilized enzyme Method comparison

"Fluorimetric Determination Of Saliva Bradykinin By HPLC With A Post-column Reaction Using Phthalaldehyde"
J. High Resolut. Chromatogr. 1986 Volume 9, Issue 5 Pages 306-307
H. Omori, N. Watanabe, T. Nakashizuka, S. Yamazaki

Abstract: A column (30 cm x 3.9 mm) packed with µBondapak C18, a mobile phase of methanol - 0.01 M sodium phosphate (pH 6.5) (3:2) at 1.5 mL min-1, and fluorimetric detection at 476 nm (excitation at 340 nm) after post-column derivatization with phthalaldehyde were used. Sample preparation was by ultra-filtration and clean-up on a Sep-Pak C18 cartridge. The calibration graph was rectilinear from 0.1 to 0.8 ng mL-1 of bradykinin, the coefficient of variation at 0.4 ng mL-1 was ~4.3%, recovery was ~85% and the detection limit (signal-to-noise ratio = 2) was 0.16 ng.
Bradykinin HPLC Fluorescence Post-column derivatization

"Sequential Flow Injection Determination Of Ionic And Total Calcium In Saliva"
J. Trace Elem. Electrolytes Health Dis. 1994 Volume 8, Issue 2 Pages 93-98
Jun S, Lima JL, Montenegro MC

Abstract: In this paper a flow injection manifold for the sequential determination of ionic and total calcium in a small (75 µL) sample of saliva is presented. This setup incorporates two detectors, a tubular potentiometric detector and an atomic absorption spectrophotometer, for determining the ionic and total calcium, respectively. Furthermore, the saliva samples can be injected directly into the manifold without any pre-treatment or loss of carbon dioxide. The results of the analyzes of 20 saliva samples were in good agreement with those obtained by the two reference procedures, the direct potentiometry for ionic calcium and atomic absorption spectroscopy for total calcium. The paired Student's t-test showed that there were no statistical differences in the results obtained. The relative standard deviations of ten consecutive measurements of the same saliva sample were approximately 3% for ionic calcium and 4% for total calcium. Effects of differences in coexisting ions, ionic strength, and pH between standard solutions and samples were negligible.
Calcium(2+) Calcium Spectrophotometry Potentiometry Method comparison Optimization