HPLC amperometry

Extraction with water immiscible organic solvent eye lens a-T, T-T RP-HPLC amperometry -I-0.8V or +0.6V 5-T 75 (1994)... 

Schematic diagrams of flow cell detectors for HPLC using (a) UVA/is absorption spectrophotometry and (b) amperometry.

Electrochemical Detectors Another common group of HPLC detectors are those based on electrochemical measurements such as amperometry, voltammetry, coulometry, and conductivity. Figure 12.29b, for example, shows an amperometric flow cell. Effluent from the column passes over the working electrode, which is held at a potential favorable for oxidizing or reducing the analytes. The potential is held constant relative to a downstream reference electrode, and the current flowing between the working and auxiliary electrodes is measured. Detection limits for amperometric electrochemical detection are 10 pg-1 ng of injected analyte. 

Detectors Most of the detectors used in HPLC also find use in capillary electrophoresis. Among the more common detectors are those based on the absorption of UV/Vis radiation, fluorescence, conductivity, amperometry, and mass spectrometry. Whenever possible, detection is done on-column before the solutes elute from the capillary tube and additional band broadening occurs. 

Specifications for modem detectors in HPLC are given by Hanai [538] and comprise spectroscopic detectors (UV, F, FUR, Raman, RID, ICP, AAS, AES), electrochemical detectors (polarography, coulometry, (pulsed) amperometry, conductivity), mass spectromet-ric and other devices (FID, ECD, ELSD, ESR, NMR). None of these detectors meets all the requirement criteria of Table 4.40. The four most commonly used HPLC detectors are UV (80%), electrochemical, fluorescence and refractive index detectors. As these detectors are several orders of magnitude less sensitive than their GC counterparts, sensor contamination is not so severe, and... 

We have already briefly described a popular application of amperometry in Chapter 13. This was the electrochemical detector used in HPLC methods. In this application, the eluting mobile phase flows across the working electrode embedded in the wall of the detector flow cell. With a constant potential applied to the electrode (one sufficient to cause oxidation or reduction of mixture components), a current is detected when a mixture component elutes. This current translates into the chromatography peak... 

All of the fat-soluble vitamins, including provitamin carotenoids, exhibit some form of electrochemical activity. Both amperometry and coulometry have been applied to electrochemical detection. In amperometric detectors, only a small proportion (usually <20%) of the electroactive solute is reduced or oxidized at the surface of a glassy carbon or similar nonporous electrode in coulometric detectors, the solute is completely reduced or oxidized within the pores of a graphite electrode. The operation of an electrochemical detector requires a semiaqueous or alcoholic mobile phase to support the electrolyte needed to conduct a current. This restricts its use to reverse-phase HPLC (but not NARP) unless the electrolyte is added postcolumn. Electrochemical detection is incompatible with NARP chromatography, because the mobile phase is insufficiently polar to dissolve the electrolyte. A stringent requirement for electrochemical detection is that the solvent delivery system be virtually pulse-free. 

The absence of a chromophore for sensitive detection is a problem which is met with HPLC determination of these thiol analytes. However, two approaches are possible to resolve this point 1) direct detection using electrochemistry, either amperometry on gold/mercury amalgamated electrodes or coulometry on porous graphite electrodes ... 

ROMANI A, MINUNNI M, MULINACCI N, PINELLI P and VINCIERI E F (2000), Comparison among differential pulse voltammetry, amperometrie biosensor, and HPLC/DAD analysis for polyphenol determination , J... 

Taha, T. S. Deits, T. L. Detection of metabolites of the Entner-Doudoroff pathway by HPLC with pulsed amperometry application to assays for pathway enzymes. Anal. Biochem. 1994, 219, 115-120. 

Detection methods for CE analysis are as diverse as those used for HPLC. The most widely used detection methods used in CE include ultraviolet/visible (UVA is) absorbance, LIE, mass spectrometry, conductivity, amperometry, radiometric, and refractive index. When deciding which detection method is best suited to an intended application, the analyst must first know if the compounds to be separated can be detected using a certain type of detector, that is, does the analyte have a chromophore. 

Electrochemical methods continue to be important as well, including inverse voltammetry, coulometry. amperometry, and potentiometry (- Analytical Voltammetry and Polarography) indeed, their overall role has actually been expanded with the development of such chemical techniques as ion chromatography and chelate HPLC. 

LC-evaporative light scattering Reverse phase HPLC-Pulsed amperometry LC-fractionation followed by NMR... 

Note A amperometry C coulometry CE capillary electrophoresis CL chemiluminescence HPLC high-performance liquid chromatography LOD detection limit MS mass spectrometry RSD relative standard deviation SF sampling frequency. 

Note A amperometry BI bead injection CL chemiluminescence F fluorescence FAT flow analysis technique HPLC/DAD high-performance liquid chromatography with diode array detector LOD detection limit MCFIA multicommutated flow injection analysis MPFS multipumping flow systems MSFIA multisyringe FIA P potentiometry RSD relative standard deviation SF sampling frequency SFA stopped-flow analysis SP spectrophotometry. 

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