What is a reference electrode? Do I need it? Which one should I use? They are the many questions that we ask ourselves!
Carbohydrates as polyalcohols can be separated at high pH using anion-exchange chromatography (HPAE). They are deprotonated, and the resulting oxoanions interact with the chosen anion exchange column. There is a whole suite of different carbohydrate columns available (Thermo Scientific Dionex CarboPac), optimized for different application needs. Under such conditions, the carbohydrates can be detected using amperometric detection. They are oxidized using a gold working electrode, and the oxidization current is used as a concentration proportional signal. To keep the detector response optimized the working electrode is cleaned by a rapid potential pulse sequence so that at the end of every fast sweep (~ 0.5 seconds) an activated fresh gold surface is available. This approach is known as Pulsed Amperometric Detection (PAD). A good overview of applications, columns, and background can be found in the Carbohydrate Testing Information.
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One of the most important, and often forgotten, parts in an HPAE-PAD instrument is the reference electrode. Its purpose is to deliver the reference potential against which the pulse sequence of PAD is delivered. The most commonly used reference electrode is the pH-electrode. In most installations, the Ag/AgCl half-cell potential serves as the reference point. Aging of pH-electrodes, whether used for PAD, or in the laboratory, is a common theme. Hence, we must regularly calibrate the pH-electrodes in the laboratory, or in PAD. In PAD, we even have to exchange it, either after six months of operation, or when the pH-readout changed more than 0.5 pH-units. You can find details in the ICS-5000+, ICS-6000, and Integrion manuals. But what if we could skip that? What if there is a reference electrode that does not need all the calibration and exchange? Sounds good? Guess how happy I was, when our colleagues published the Technical Note 73348 titled “Carbohydrate determinations by HPAE-PAD using a PdH reference electrode”. Manali and Jeffrey compared the use of both reference-electrode-type running typical carbohydrate separations. It is quite an interesting read, and worth a thought when you need to exchange the reference electrode the next time.