Quantification of Aflatoxins B₁, B₂, G₁, and G₂ in Pepper by HPLC/Fluorescence

Abstract

Sample cleanup with Sep‐Pak cartridges and validation of an HPLC/fluorescence method for separation and identification of aflatoxins in pepper was carried out. The samples were extracted with chloroform/water and filtered. The filtrate was purified on a silica cartridge followed by a C₁₈ cartridge. Aflatoxins were derivatized with trifluoracetic acid and analyzed by HPLC with a column containing TSK gel ODS 80 TM CTR. Validation of the method was evaluated by recovery studies and CV%. Samples of black and white pepper were spiked in triplicate with total aflatoxins B₁, B₂, G₁, and G₂ at 3, 6, and 12 µg/kg levels. Recovery values were in the range of 86.2–96.2 for aflatoxin B₁, 91.2–97.7 for aflatoxin B₂, 85.3–94.2 for aflatoxin G₁, and 89.8–97 for aflatoxin G₂ with CV% being less than 12.1%. The detection limits for each aflatoxin, based on a signal‐to‐noise ratio of 3, were 0.009 µgL⁻¹ for G₁ and 0.006 µgL⁻¹ for B₁, G₂, and B₂. Twenty‐two samples of pepper (4 black and 18 white) were analyzed for aflatoxins by high performance liquid chromatography with fluorescence detection. The degree of contamination in a lot of white pepper imported from India (six samples) and in sixteen samples randomly purchased from the market was estimated. Fourteen of the analyzed samples contained aflatoxins (B₁, B₂, G₁, or G₂); however, they were below the limit of 20 ppb fixed by the European Union, except for one.

Keywords:

• Aflatoxins
• Pepper
• HPLC/fluorescence