University of North Florida
Browse the Citations
-OR-

Contact Info

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

View Stuart Chalk's profile on LinkedIn

Stephen G. Weber

Abbrev:
Weber, S.G.
Other Names:
Address:
Departemnt of Chemistry, University of Pittsburgh, Pittsburgh, PA 15260, USA
Phone:
NA
Fax:
NA

Citations 15

"Cyclic Voltammetry Of S-sulfocysteine At A Gold Plus Mercury Amalgam Electrode And Application To Dual Electrode Electrochemical Detection"
J. Electroanal. Chem. 1997 Volume 435, Issue 1-2 Pages 39-46
Wanlin Xiaa, Steven J. Woltmana, Mats Sandbergb and Stephen G. Webera,*

Abstract: The dual gold + mercury (Au + Hg) amalgam electrode was found to be an effective detector for S-sulfocysteine (SSC). Using flow injection analysis, a 20 pmol mass detection limit was achieved using a 20 µL injection volume. Cystine and SSC were shown to behave quite differently in this detection system. The signal from cystine displayed a dependence on flow rate reflective of simple mass transfer, while the signal from SSC was more sensitive to mobile phase flow rate. The cystine signal was insensitive to pH, while SSC's signal depended significantly on pH. The SSC reduction reaction at the upstream Au + Hg electrode was determined by cyclic voltammetry to be RSSO3- + 2H(+) + 2e(-) --> RSH + HSO3-. The signal monitored at the downstream electrode was attributed mainly to the oxidation of Hg to Hg cysteinate below pH 4. Both cysteine and bisulfite contributed to the detection signal above pH 4.5. The optimum pH for the detection of SSC was approximately 5.2. (C) 1997 Elsevier Science S.A. 22 References
S-sulfocysteine Electrode Voltammetry Electrode Optimization

"Simple Method For The Quantitative Examination Of Extra Column Band Broadening In Microchromatographic Systems"
J. Chromatogr. A 2003 Volume 986, Issue 2 Pages 247-251
Amy T. Beisler, Kathleen E. Schaefer and Stephen G. Weber

Abstract: In recent years capillary chromatography has gained popularity for trace analyzes. Most often UV or electrochemical detection is employed because the small peak volumes make post-column derivatization challenging. We have developed a simple method based on flow injection for determining contributions to peak broadening from post-column reactors. The only requirement for application of our methodology is that diffusion be in the Taylor regime so that radial concentration gradients are relaxed enabling mixing purely by diffusion.

"Optimization Of A Modified Electrode For The Sensitive And Selective Detection Of α-dipeptides"
J. Chromatogr. A 1995 Volume 705, Issue 2 Pages 171-184
Jian-Ge Chen, Edward Vinski, Kevin Colizza and Stephen G. Weber*

Abstract: The modified electrode was prepared by subjecting a vitreous C electrode to 1.2 V vs. Ag/AgCl for 4 h in a flowing electrolyte (1 ml/min) containing 0.1 mM Cu in phosphate buffer of pH 8. The selectivity of the modified electrode was altered by controlling the pH and composition of the detection eluent. The electrode was more selective for α-dipeptides over β- and γ-dipeptides and amino-acids at pH 9.8. At pH 8, the electrode was selective for all dipeptides over amino-acids. The detection limits for dipeptides were ~e;10 nM in a flow injection system employing a 20 µL sample loop, a detection potential of 1.1 V vs. Ag/AgCl and an eluent (1 ml/min) containing 0.1 mM Cu in phosphate buffer of pH 8. Surface analysis showed that the oxidation of the vitreous C electrode gave rise to its selectivity. The oxidation of dipeptides at the modified electrode was completely inhibited by 10 mM Mg2+ in the eluent. The modified electrode was applied to the detection of eight dipeptides and glutathione (four α-, two β- and two γ-dipeptides and γ-glutamyl tripeptide) following separation by anion-exchange chromatography (details given). Sensitive and selective detection of dipeptides is important in neurochemistry. We have developed a flexible detection scheme for dipeptides based on a modified carbon electrode. The modification arises from the anodic treatment of the carbon electrode in alkaline solution. The flexibility of the detection scheme arises from the different conditions used in both the modification and the detection. It is shown that the modification step requires the presence of cupric ion, while the detection step does not. On the other hand, it is shown that the presence of copper in the detection eluent, as well as the pH of the environment, can be used in controlling the selectivity of the modified electrode. For example, the modified electrode is more selective for α-dipeptides over β- and γ-dipeptides as well as amino acids at pH 9.8, whereas it is selective for all dipeptides over amino acids at pH 8.0. Detection limits of dipeptides on the order of 10 nM were achieved at pH 8.0 by flow injection analysis with a knotted Teflon tubing connecting the injector and the detector that gave a typical peak volume of about 0.50 mL at 1.0 ml/min. From surface analysis it is shown that the oxygenation of the glassy carbon electrode gives rise to the selectivity. The oxidation of dipeptides at the modified electrode is completely inhibited by 10 mM Mg2+ in the eluent.
Disaccharides Glutathione Electrode Interferences Mixing coil Knotted reactor Optimization

"Electrochemical Detection Of Oligopeptides Through The Pre-column Formation Of Biuret Complexes"
J. Chromatogr. A 1991 Volume 542, Issue 2 Pages 345-350
Hweiyan Tsai and Stephen G. Weber

Abstract: The post-column reaction for the formation of electroactive Cu(II) - peptide complexes from larger (>6 amino-acids) peptides requires relatively high temperature and a long reaction time. As an alternative, the incubation of three larger peptides (bradykinin, Tyr8-bradykinin and insulin A chain) with biuret reagent for 20 min at 60°C yields complexes which can be analyzed by flow injection analysis or LC using acidic or basic mobile phases and electrochemical detection. The sensitivity of the latter is around 1.0 nC pmol-1 at 0.80 V and a flow rate of 1 mL min-1. The influence of the column packing material on the electrochemical detector response of the Cu - peptide complexes is also considered.
Oligopeptides LC Electrochemical analysis Heated reaction

"Voltammetry In Static And Flowing Solutions With A Large-amplitude Sine Wave Potential"
Electroanalysis 1992 Volume 4, Issue 4 Pages 429-437
John T. Long, Stephen G. Weber *

Abstract: Voltammetric detection is a potentially powerful method of detection for high performance liquid chromatography and flow injection analysis. A problem that needs a solution is its inferior detection limit compared to that obtained with amperometric detection. The shape of the potential waveform has been investigated in the search for methods of improving the detection limit of voltammetric detection. As a result, large amplitude sine wave potential perturbations have been found to give improved signal-to-noise ratios when compared to the values found for a corresponding triangle waveform (by a factor of from 1.1 to 5). This is principally due to the fact that the background current from a large amplitude triangle perturbation is spread over a wider frequency region. Signal processing techniques such as digital filtering have also been investigated. Digital bandpass filtering can be used to improve the signal to noise ratio by a factor of 2-13. Qualitative information (e.g., half-wave potential) is obtained through observing the phase angle at harmonics of the fundamental frequency.

"Tris(2,2'-bipyridine)ruthenium (II) As A Peroxide-producing Replacement For Enzymes As Chemical Labels"
Biosens. Bioelectron. 1991 Volume 6, Issue 8 Pages 699-705
Kamal Z. Ismail and Stephen G. Weber*

Abstract: The concentration of the ruthenium-based label is determined from the rate of hydrogen peroxide production elicited by photolysis. Electron transfer quenching of the photoexcited label by methyl viologen (1,1'-dimethyl-4,4'bipyridiniu m dication, MV2+) and/or oxygen in the presence of EDTA generates hydrogen peroxide. Both flow injection and direct photolysis techniques were tested, with the latter showing better results. Direct photolysis is more sensitive, faster, requires only a 20 µL sample volume, uses only 30 mW laser power and shows a smaller background. The presence of 5% normal human serum in the sample did not interfere with the measurements. Linear calibration curves were obtained in the nanomolar concentration range for goat antimouse antibody labeled with the ruthenium complex. The determination of membrane-surface-bound labeled IgG is accomplished by direct photolysis of a membrane that covers a platinum microelectrode.
Ruthenium(II) Electrode Spectroelectrochemistry Enzyme Interferences Quenching Sensitivity Laser

"Analysis Of The Performance Of A Flow Reactor For Use With Microcolumn HPLC"
Anal. Chem. 2004 Volume 76, Issue 3 Pages 639-645
Amy T. Beisler, Eskil Sahlin, Kathleen E. Schaefer and Stephen G. Weber

Abstract: Post-column derivatization reactions can be used to improve detector sensitivity or selectivity. The advantages of capillary chromatography for trace analysis could be augmented if there were post-column reactors suitable for micro-chromatographic systems. However, post-column derivatization is a challenge because of the small peak volumes associated with capillary columns. We have developed a post-column flow reactor from micro-channels formed in fluorinated ethylene propylene and 50-µm fused-silica tubing for use with capillary HPLC analyzes. Theoretical and experimental evidence show that the reactor, which operates in the Taylor dispersion regime, enables contact of analyte and derivatization streams purely by diffusion. Reactor lengths as short as 2 cm allow formation of copper(II)-peptide complexes that are detected electrochemically at a carbon fiber micro-electrode. The reactor has been used with 100 µm i.d. columns with insignificant effects (i.e., <3%) on peak band spreading. Theoretical calculations indicate that even smaller i.d. columns can be used with little effect on chromatographic resolution.

"Miniaturized Electrochemical Flow Cells"
Anal. Chem. 2003 Volume 75, Issue 4 Pages 1031-1036
Eskil Sahlin, Alexandra ter Halle, Kathleen Schaefer, Jeffery Horn, Matthew Then and Stephen G. Weber

Abstract: Several novel types of miniaturized electrochemical flow cells are described. The flow cells are fabricated in fluorinated ethylene propylene using a novel technique where channels with inner diameters down to 13 µm are integrated with electrodes. The channel is formed by shrinking and simultaneous melting of a heat shrink/melt tubing around a channel template (a tungsten wire) and electrodes followed by removal of the channel template. The technique allows incorporation of different electrode materials of different sizes. The electrode configuration consists of one or two working electrodes inside the channel and a counter electrode located in the channel outlet reservoir. Electrode configurations with different channel and working electrode sizes, different electrode materials including carbon fibers, glassy carbon rods, poly(tetrafluoroethylene)/carbon composite material, and platinum wires, and different arrangements have been assembled. Hydrodynamic voltammograms in dual-electrode (generator-collector) experiments indicate good potential control for cells with 25-µm channels, while there is some iR drop in cells with 13-µm channels. Cells prepared with a cylindrical working electrode tangent and perpendicular to a flow channel show a flow rate dependence consistent with thin-layer cell behavior. Electrode areas can be made in the range of 10^-10-10^-8 m2.

"Chromatographic Detection Using Tris(2,2-bipyridyl)ruthenium(III) As A Fluorogenic Electron-transfer Reagent"
Anal. Chem. 1999 Volume 71, Issue 8 Pages 1504-1512
Steven J. Woltman, William R. Even, and Stephen G. Weber

Abstract: Tris(2,2-bipyridyl)ruthenium can be excited to fluorescence by visible light (λabs 454 nm, λem 607 nm) when in the M(II) oxidation state, but not in the M(III) state. A novel chromatography detection method using the nonfluorescent M(III) form of the complex as a post-column fluorogenic reagent is demonstrated. The M(III) form is a powerful oxidizing agent (E°C = 1.27 V vs. NHE, 1.05 V vs. Ag/AgCl). The M(III) reagent is generated online from the M(II) form of the complex by a highly efficient porous carbon electrode and then reacted briefly with chromatography effluent; the M(II) created by electron transfer from oxidation-susceptible analytes is then detected by fluorescence. The fluorescence detector can be calibrated for number of electrons transferred by injection of either M(II) or an oxidative standard such as ferrocyanide. It is hoped that this redox-based detection scheme will provide an alternative to electrochemical detection. Among the advantages are freedom from surface fouling and the potential for extremely low detection limits. The scheme was applied to detection of the peptide dynorphin A and several of its fragments. Dynorphin A contains tyrosine at the N-terminus (position 1) and tryptophan in position 15; these amino acid residues are susceptible to oxidation and peptides containing them can be detected on that basis. Flow injection testing of the model compounds Tyr-Gly-Gly-Phe-Leu and Gly-Gly-Trp-Gly indicated that tyrosine transferred ~1 electron to the M(III) reagent and that tryptophan transferred ~4 electrons. Similar results were obtained from the chromatography runs. Dynorphin A and six dynorphin A fragments containing the N-terminal tyrosine were detected easily at 100 nM concentration. (14 pmol) using laser-induced fluorescence. As expected, one fragment that did not contain tryptophan or tyrosine was not detected. A mass detection limit of 80 fmol was estimated for the tyrosine-containing fragments.

"Optical Control Of Divalent Metal Ion Binding To A Photochromic Catechol: Photoreversal Of Tightly Bound Zn2+"
Anal. Chem. 1999 Volume 71, Issue 6 Pages 1146-1151
Mark T. Stauffer and Stephen G. Weber

Abstract: Photochromic molecules with functional groups that bind metal ions strongly make possible the development of photoreversible sensors and pre-concentrators for detection of metal ions at trace concentrations. While several photochromic metal chelators have been reported, none have been shown to bind strongly to divalent metal ions. A novel photochromic benzopyran 2, which is a photochromic catechol, binds Zn2+ strongly and reversibly. The thermal equilibrium between open and closed 2 is slow. Thus, binding of Zn2+ to 2 is slow in the dark. Under illumination with 306-416-nm light, Zn2+ binds rapidly to open 2. The stable complex dissociates upon illumination with visible light, showing that both formation and dissociation of the complex are controllable with light. A binding constant of 1.1 x 10(9) is estimated from a Job plot, which also indicates formation of a 1:1 Zn2+/open 2 complex.
Chelation

"Photoelectrochemical Sensor For Catalase Activity Based On The In Situ Generation And Detection Of Substrate"
Anal. Chem. 1993 Volume 65, Issue 2 Pages 169-175
Claudia B. Cohen and Stephen G. Weber

Abstract: A device for the in situ generation and detection of hydrogen peroxide is presented for application to the automation of enzyme immunoassay and application to biosensors. The device is fabricated from a gold-coated optical fiber. It photochemically generates and electrochemically detects H-2O-2 in aqueous, buffered solutions. The electrochemical signal is attenuated in the presence of the enzyme catalase (EC 1.11.1.6), for which H-2O-2 is a substrate. The quantum efficiency of the peroxide-producing reaction (0.20%), the collection efficiency of the gold ring electrode ((3.8 +- 0.05) times 10^-3), and the sensitivity of the photocurrent to catalase have been evaluated. The sensor's photosignal is reduced by 50% in the presence of 25 nM catalase. Detection limits of 76 pM catalase have been obtained using FIA.
Enzyme, catalase Immunoassay Sensor Optical fiber Optimization Detection limit

"Influence Of Tyrosine On The Dual Electrode Electrochemical Detection Of Copper(II) - Peptide Complexes"
Anal. Chem. 1992 Volume 64, Issue 23 Pages 2897-2903
Hweiyan Tsai and Stephen G. Weber

Abstract: A study has been made of the effect of tyrosine on the determination of peptides by LC with detection as the Cu(II) complexes (biuret reaction). Samples were analyzed (i) on a column (25 cm x 4.6 mm) of Biotage PBD with a mobile phase containing 0.1 mM CuSO4, 0.3 mM K Na tartrate, 0.1 M NaOH and 0.2 M NaHCO3 (pH 9.80 to 9.85), or (ii) on a column (15 cm x 2 mm) of Waters Delta Pak C18 with 0.1% of trifluoroacetic acid in aqueous 10% acetonitrile as mobile phase and, as post-column reagent, the mobile phase from (i). The detector was a BAS dual vitreous carbon electrode with a model LC-4B potentiostat to control the potential (vs. Ag - AgCl, 3 M NaCl) and measure the anodic current. Flow injection analysis was performed with the mobile phase as in (i). Peptides were separated in system (ii) but not in system (i). The presence of tyrosine in the peptides increased the anodic sensitivity to the Cu(II) - peptide complexes by a factor of two to four, affording an anodic detection limit of 40 to 100 fmol, but impaired the cathodic sensitivity; the effect of tyrosine depended on its position in the peptide molecule.
Copper(II) Electrode Interferences Complexation

"Recovery Of Voltammograms By Target Factor Analysis Of Current - Time Data In Electrochemical Detection"
Anal. Chem. 1990 Volume 62, Issue 23 Pages 2643-2646
Gregor Reich, Josef Wolf, John T. Long, and Stephen G. Weber

Abstract: The use was studied of target factor analysis (TFA; Malinowski and Howery, Factor Analysis in Chemistry, Wiley, New York, 1980) in the removal of noise from voltammograms obtained from 0.15 µM to 0.15 mM adrenaline and 0.14 µM to 0.14 mM tris-(2,2'-bipyridyl)iron perchlorate solution in a flow injection analysis system with voltammetric detection using a sinusoidal potential waveform. A modified version of the FACTANAL program (QCPE Program No. 320, Indiana Univ., Bloomington, IN) was applied to the experimental data and results were compared with those of classical background noise subtraction method. The signal-to-noise ratio requirements necessary for successful TFA, and the effects of a combination of a stationary and a changing background on TFA are discussed.
Electrochemical analysis Voltammetry Signal to noise ratio

"Detection Limits And Selectivity In Electrochemical Detectors"
Anal. Chem. 1988 Volume 60, Issue 15 Pages 903A-913A
Stephen G. Weber, John T. Long

Abstract: Developments in current-carrying electrochemical detectors for use in flow, e.g., liquid chromatographic, systems are described and reviewed with special reference to signal and noise generation, signal-to-noise ratio, improvement of qualitative information content and control of selectivity. Future developments in the versatility and potency of electrochemical detection are discussed. (53 references).
Electrochemical analysis Review

"Theoretical And Practical Limitations On The Optimization Of Amperometric Detectors"
Anal. Chem. 1984 Volume 56, Issue 6 Pages 978-985
Janean M. Elbicki, Donald M. Morgan, and Stephen G. Weber

Abstract: Steady-state equations for hydrodynamic current in amperometric detectors, when integrated, may be used to characterize detectors in flow injection systems, e.g., in liquid chromatography. Peak areas are independent of peak shape. Theory can be correlated in this way with experimental results. Optimization can then be considered by examination of the graphical region where five inequalities derived from boundary conditions are satisfied. Wall-jet and channel thin-layer cells have been studied in detail and found to be equivalent when small spacers are used. The dependence of current on flow rate lessens with time due to the formation of a thin film on the electrode surface. Graphs are presented of the inequalities for both thin-layer and wall-jet cells.
HPLC Amperometry Electrode Optimization Peak shape