Reference Electrodes

Different Reference Electrodes

By far the most familiar reference electrode to electrochemists is the standard hydrogen electrode (SHE), from which many standard half-reaction redox potentials have been obtained and are to be found in many reference handbooks. The SHE is far too impractical for routine HPLC-ECD requiring hydrogen gas, so several other reference materials have been developed and are now in use.

The silver/silver chloride reference is a "wet" electrode of the metal/insoluble salt type. It is usually placed downstream from the working electrode and makes "electrical" contact with the mobile phase via a microporous plug made from materials such as ceramic or Vicor®. Although more practical to use than the SHE, its relatively large size necessitates its placement far away from the working electrode. Such a configuration can lead to multiple junction potentials and iR drops, which ultimately lead to excessive noise, instability of the applied potential, and a nonlinear response. This reference electrode requires routine maintenance, is photosensitive, can be readily poisoned by sulfide and other anions, and may become blocked at the microporous plug.

Recently, Bohs et al. (1989) used a solid silver wire coated with silver chloride as a reference electrode. Although the reference can be placed close to the working electrode to overcome issues with junction potentials and iR drops, the electrode design necessitates the use of 10mM sodium chloride in the mobile phase. The reference is fairly unstable, capable of altering its reference potential by 60mV/month and shifting its potential (resulting in prolonged voids, system peaks and unreliable quantitation) when exposed to solutions high in chloride ions - as with the analysis of microdialysis perfusates. The incorporation of 10mM sodium chloride into the mobile phase may diminish such problems but also may contribute to system noise, system corrosion and high background currents.

A similar reference electrode to the wet silver/silver chloride electrode is the mercury/mercurous chloride or calomel electrode. The construction of this electrode is more complex than the silver/silver chloride and suffers many of the same problems. In addition, it uses highly toxic salts.

Both the wet silver/silver chloride and calomel electrodes must be placed in the HPLC system in which the back pressure is relatively low - usually post column and downstream of the working electrode. Failure to do so may force mobile phase into the reference compartment through the porous plug, possibly rupturing the reference electrode or changing the reference potential. Electrodes designed to remove electroactive interferences from the mobile phase, such as the ESA Guard Cell (Model 5020), must be able to withstand high back pressures, as they are typically placed before the column. Furthermore, when multiple electrodes are placed in series to form an electrode array, the back pressure on the upstream electrodes can be fairly high. In both cases the analytical electrodes cannot use either the wet silver/silver chloride or calomel references.

Probably the most useful reference electrode is the a-hydrogen/palladium electrode (Matson, 1983). This reference is very small and can be placed in close proximity to the working and counter electrodes, thereby overcoming possible iR drops.¹ This reference electrode is used in all ESA analytical sensors including the flow-through porous graphite coulometric sensors as well as the thin-layer amperometric sensors. It is maintenance free, stable, not easily poisoned and capable of working at high pressures (>6000 psi). Furthermore, it is pH sensitive. The reference electrode shifts about 60 mV/pH unit. The changes in the redox potential of an analyte as a result of a change in pH can therefore be compensated for by an accompanying shift in the reference potential. This approach overcomes significant shifts in analyte response due to pH shifts that are common with thin-layer detectors. Furthermore, it can be used with pH gradients and, unlike the silver/silver chloride reference, is unaffected by major perturbations in the baseline. The potential of the Pd wire has been ascribed to be a mixed potential effect (Hoare, 1964: Vracar, et al., 1987; Shao, et al., 1994). Many analytes are also pH sensitive (e.g., catechols) and any pH drift in mobile phase can be compensated for by an equal shift in reference potential.

¹ iR is important because if the iR is large (high current and/or high resistance) the potential shift and analysis could be non-linear, at least at high analyte concentrations.