Applied Potential at the Reference Electrode

Importance of the Reference Electrode

A common practical problem occurs as a result of misunderstanding the importance of the reference electrode. Often a method developed on one particular HPLC-ECD system does not work well when transferred to another system using a different reference electrode. For example, catecholamines can be readily measured at +800mV against a silver/silver chloride reference but the same potential used with a palladium reference, might cause the assay to fail from excessive noise and interferences, resulting in poor limits of detection. There is a difference of approximately 300mV between the silver/silver chloride and palladium reference electrodes, which means that if the working electrode was set to +800mV against a palladium reference then this would be equivalent to +1100mV vs. silver/silver chloride! Readers are strongly urged to check the reference type when trying published methods in their laboratory, produce an HDV, and then adjust the working potential of their system accordingly. If the applied potential using a silver/silver chloride reference electrode is set to +500mV, this is equivalent to an applied potential of +200mV using a Coulometric electrode with its palladium reference.

When a potential is first applied to the working electrode (or the potential is changed), charging currents are produced at the solution/electrode interface. Charging current peaks and decays exponentially. The time to reach zero can be on the order of hours as with many thin-layer types with silver/silver chloride reference designs, meaning considerable analytical down-time will occur if this detector is turned off or the potential is changed. With the coulometric electrode, such charging currents decay more rapidly and can reach zero in only a few minutes. It is common to prolong the porous graphite electrode life by applying a potential difference of +1000mV to the working electrodes for 5 minutes to electrochemically clean the working electrode surface. High-sensitivity analyses can be restarted only minutes after the potential change.

Summary

A variety of reference electrodes exist for use in HPLC-ECD.  The wet silver/silver chloride electrode was historically one of the first reference electrodes to be used with HPLC-ECD. However, it is unstable, requires maintenance, is affected by both pH and light, and can readily be poisoned. It is relatively large and cannot be placed in close proximity to the working electrode which may result in iR drops and problems with the detector. Furthermore, due to a major limitation in its design, a wet electrode cannot be subjected to pressure.

The -hydrogen/palladium reference electrode is superior to most other reference electrodes. It is maintenance free, unaffected by light and not easily poisoned. It is small and can be placed close to the working electrode for better potential control. It is pH sensitive, therefore the changes in the redox potential of an analyte, resulting from a change in pH, are compensated for by an accompanying shift in the reference potential. Finally, this reference electrode is unaffected by changes in pressure. This reference electrode is used in all ESA electrochemical cells.