Hot Pepper Compounds Caught by Cool Coulometric Detection

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  • Published: Apr 17, 2017
  • Author: Ryan De Vooght-Johnson
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thumbnail image: Hot Pepper Compounds Caught by Cool Coulometric Detection

Objective measurement of pepper heat required

Traditionally, the determination of the ‘hotness’ of peppers and derived foodstuffs is measured on the ‘Scoville scale’, which involves a taster detecting the ‘hot’ taste through successive dilutions. Such scoring is subjective, with the results varying between tasters, so the food industry requires more objective methods for determining the heat of products.

A group of compounds known as capsaicinoids is responsible for the heat. They have been detected by GC, capillary electrophoresis and HPLC, with UV, mass spectrometry, amperometric or fluorescence detection having been combined with the latter technique. Some of these methods lack the sensitivity to detect traces of capsaicinoids, which may still give some flavour sensation. Isolation of capsaicinoids from various food matrices can also be a challenge. Since capsaicinoids are phenolic compounds, a coulometric detection method can be employed in conjunction with HPLC. Scientists from the Free University of Brussels, Tokyo Kasei University and the College of Analytical Chemistry in Osaka have devised a novel HPLC method with coulometric detection to assay capsaicinoids.

HPLC with coulometric detection used for capsaicinoids

Samples of sliced peppers and different types of seasoning containing pepper were extracted with methanol under sonication. A solid-phase extraction (SPE) cartridge (Intersep GL Pak GC/NH2) was used to extract the capsaicinoids from the methanol solution. The compounds were eluted from the cartridge with acetone:methanol (7:3 v/v). Filtration and evaporation gave a concentrate of the desired compounds, which was taken up in methanol and then diluted with mobile phase prior to injection onto the HPLC.

The three main capsaicinoid compounds, capsaicin, dihydrocapsaicin and nordihydrocapsaicin, were detected by HPLC using a Showa Denko Shodex C18M 4D column. The mobile phase was a 51:49 mixture of acetonitrile and 0.15 M acetate buffer at pH 3.5, with a 0.8 ml/min flow rate. An ESA Coulochem coulometric detector was used; this contained an initial ‘guard cell’, which oxidised unwanted species that would interfere with the capsaicinoid peaks, followed by an analysis cell. The potential of the guard cell was set to be lower than that of the analysis cell, so that it would not oxidise the capsaicinoids. The analytical cell (Model 5010) consisted of a porous carbon working electrode, a platinum counter electrode and a palladium reference electrode, with the potential being set at 0.65 V vs. Pd.

The three peaks were clearly separated, although the peaks from capsaicin and nordihydrocapsaicin were close, and it was found to be important to have the correct solvent composition in order to obtain separation. The system gave good linearity and a limit of detection (LOD) of only 10 pg per injection, which is comparable to LC-MS and amperometric detection.

The results showed wide variations in the amount of the three capsaicinoids between different types of pepper and seasoning products. As expected, the habanero pepper gave the highest readings, with 240 μg/g of product for capsaicin, 149 μg/g dihydrocapsaicin and 32 μg/g nordihydrocapsaicin, while the corresponding figures for a green pepper were 0.16 μg/g, 0.41 μg/g and none detected.

Coulometric cells give good HPLC results for capsaicinoids

The paper has shown that good results for capsaicinoid detection can be obtained using the SPE isolation method and coulometric HPLC. The quantification of these compounds removes the need for food companies to rely upon the vagaries of subjective tests. Coulometric methods are seldom used for routine analysis, since UV or MS detection are normally employed with HPLC. However, electrochemical cells can give excellent results for compounds that they can readily oxidise or reduce, such as phenols or hydroperoxides. Such techniques could therefore become more widely used in the future.

Related Links

Electroanalysis, 2017, Early View paper. Watanabe et al. Determination of three capsaicinoids in raw red peppers and seasoning powders by liquid chromatography with coulometric detection.

Free Radical Biology and Medicine, 1996, 20, 365-371. Arai et al. Coulometric detection in high-performance liquid chromatographic analysis of cholesteryl ester hydroperoxides.

Wikipedia, Capsaicin

Article by Ryan De Vooght-Johnson

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

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