Reuse of regenerated immunoaffinity column for excellent clean-up and low-cost detection of trace aflatoxins in malt

Abstract

The high price of the immunoaffinity column (IAC) has limited its broad application in the cleanup of analytes from complex matrices. This study aimed to reduce practical cost by regenerating IACs for reuse in cleaning up and purifying aflatoxin (AF) B1, B2, G1 and G2 from medicinal and edible malt samples, followed by high-performance liquid chromatography coupled with photochemical derivatization and fluorescence detection (HPLC-PCD-FLD). The IACs were prepared in advance by coupling anti-AFs specific antibodies (Abs) on a solid phase carrier. After each use, the IACs were washed immediately with phosphate-buffered saline (PBS) and pure water, respectively, and then were filled with PBS as the preservation solvent for storage at 4 °C overnight to recover the binding activity of the immobilized Abs for the regeneration and reuse in the next day until the recovery rate was lower than 70%. Results showed that the developed HPLC-PCD-FLD method displayed good linearity in wide concentration range with the limit of detection (LOD) and quantification (LOQ) of 0.3125–0.5 ng/mL and 1–1.5 ng/mL for the four aflatoxins, as well as excellent accuracy and precision with relative standard deviations (RSDs) <5.5% and <7.0%, respectively. The IACs could be used at least nine times for AFB2, AFG1, and AFG2 when intruding the recommended method in Huaan Magnech Bio-Tech IAC instructions for raw malt sample preparation. And the corresponding consumption could be reduced to one-ninth of the original cost. Due to the high toxicity and incidence of AFB1, the IACs were suggested for reuse twice regarding this aflatoxin in malt with half of the original cost. The reuse of the regenerated IACs for excellent clean-up and low-cost detection of trace AFs in complex matrices is economical, green and environment-friendly, expecting widespread application of more mycotoxins in sophisticated food and traditional Chinese medicine matrices with low cost.