Rapid Determination of Diarrhetic Shellfish Poisoning in Shellfish by Colorimetric Protein Phosphatase Inhibition Assay
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Abstract:
Okadaic acid (OA) and its derivatives, the components of diarrhetic shellfish poisoning (DSP), are potent inhibitors of protein phosphatases. Based on the mechanism of action, a protein phosphatase 2A (PP2A) inhibition assay for the rapid determination of DSP toxins in bivalve was developed. In this study, p-nitrophenyl phosphate (p-NPP) was used as the substrate and hydrolyzed by PP2A, then the product was measured at 405 nm. The total DSP content (calculated by OA) in samples could be detected according to the standard dose-effect curve developed with a series of OA standard solutions. The experimental conditions of sample preparation were optimized, and shellfish matrix loading limits for the protein phosphatase inhibition assay were established according to the shellfish species. The detection limit was 80 μg/kg, andthe spiked recoveries for OA in shellfish samples were between 90.43% and 118.52%, with relative standard deviations (RSD) ranged from 6.85% to 13.93%. The colorimetric protein phosphatase inhibition assay was simple, rapid and showed good recovery and reproducibility, demonstrating this proposed method could be used as an efficient analysis tool for rapid screening of DSP in shellfish and suit for daily monitoring to control shellfish toxicity.
