Cheap, disposable but effective: a disposable amperometric detector for HPLC

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Ezine

  • Published: Jun 20, 2011
  • Author: Jon Evans
  • Channels: Detectors
thumbnail image: Cheap, disposable but effective: a disposable amperometric detector for HPLC

Scraping clean

Amperometric detection, in which analytes are detected based on the current they generate when oxidised or reduced at an electrode, has several important advantages as a detection technique. It can detect a wide range of compounds, especially organic compounds, without requiring them to be chemically modified in any way; it's also fairly small-scale, requiring little more than a few electrodes, and is thus ideal for incorporation onto lab-on-a-chip systems.

Unfortunately, it also has one major drawback, which, even more unfortunately, is inherent to the way it works. Oxidizing or reducing analytes at an electrode, termed the working electrode, tends to coat it with residue, eventually stopping it from working properly. As a consequence, the working electrodes in amperometric detectors regularly need to be scraped clean, which is a fairly time-consuming process.

An alternative approach is simply to throw the detector away when the working electrode becomes unusable and replace it with a new one, which is fine as long as you have a cheap, disposable amperometric detector. This is what a team of US chemists led by William Heineman at the University of Cincinnati in Ohio has now developed for high performance liquid chromatography (HPLC).


Tiny electrodes

Their detector consists of a microfluidic chip just 5cm long and 2.5cm wide containing a network of microscopic channels. This network includes four sample channels, one channel for the mobile phase and two waste channels. The four sample channels all culminate in a flow cell region containing three tiny electrodes - a working electrode, an auxiliary electrode and a reference electrode - for amperometric detection.

The working and auxiliary electrodes, which between them generate the electric potential that instigates the oxidation and reduction reactions, are made of platinum and are each just 0.17mm2 in size. The reference electrode, against which the fluctuating current generated at the working electrode is compared, is even smaller, at just 0.031mm2, and is made of platinum coated with silver and silver chloride.

Samples and a mobile phase can be pumped directly onto the microfluidic chip, allowing it to be used for flow injection analysis (FIA), or the chip can be hooked up to an HPLC column to act purely as an amperometric detector. As a first test of the chip, Heineman and his colleagues used it for FIA, finding that it could repeatedly detect potassium ferricyanide.

Next, they used it with HPLC, finding that the chip could detect ascorbic acid (vitamin C) and acetaminophen (paracetamol) after they had been separated, as well as the structurally similar pharmaceutical compounds dextrorphan and levallorphan, even when spiked into human plasma.


Cost cutting

Compared to a commercially-available amperometric detector, the microfluidic chip was much less sensitive, with limits of detection for dextrorphan and levallorphan in human plasma of 50nM and 80nM, respectively, compared to 10nM and 20nM for the commercially-available detector. The main reason for this is that the commercially-available detector is much bigger, with electrodes that are almost 300 times larger than those used on the microfluidic chip. Nevertheless, these limits of detection are small enough for many applications and the microfluidic chip also produced a highly linear response to varying concentrations of the analytes.

When used with HPLC, the microfluidic chip began to stop working properly after a maximum of around 20 hours. Rather ironically, this wasn't because the working electrode became coated with residue, but rather because the flow of liquid through the channels washed away the silver coating from the reference electrode. So now Heineman and his team are working on ways to protect this coating or make it more robust, thereby ensuring that only the working electrode stops the chip from working properly.

They are also looking at reducing the cost of the chip. At the moment, they estimate that 100,000 of these chips could be produced for around $25 each. But by redesigning the chip so that only the electrode section needs to be replaced, Heineman thinks this could be reduced to less than $15.



The views represented in this article are solely those of the author and do not necessarily represent those of John Wiley and Sons, Ltd.

Cheap, disposable but effective: a disposable amperometric detector for HPLC

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