A portion of 5 g from each subsample of hot pepper was oven-dried at 60 °C for 2–5 days, cooled and then ground to dried pepper powder. Samples were analyzed by using a chromatographic method previously described . A mixture of 1-g sample with 10 mL acetonitrile was placed in 120-mL glass bottles with Teflon-lined lids, capped and placed in a water bath at 80 °C for 4 h and swirled manually every hour. The bottles were removed from the water bath and cooled at room temperature. The supernatant content of samples (2–3 mL) was filtered through a 0.45 μm filter (Millex®-HV filter) using a 5-mL disposable syringe (Millipore, Bedford, MA, USA) into an HPLC sample vial. For the liquid chromatographic analysis of capsaicinoids, an HPLC system LC-10 (Shimadzu, Kyoto, Japan) equipped with an SPD-10A. UV-Vis detector (set at 280 nm wavelength) was used. The analysis was carried out with the isocratic mobile phase (acetonitrile:water, 60:40) at a flow rate of 1 mL/min using a column Discovery C18 (250 × 4.6 mm, 5 μm) supplied by Supelco (Bellefonte, PA, USA). The limit of detection (LOD) was 0.1 μg/g for both capsaicinoids.