Determination of Paralytic Shellfish Poisoning Toxins by Liquid Chromatography with Fluorescence Detection Using Pre-column Derivatization with Hydrogen Peroxide Oxidation

Saxitoxin (STX) and decarbamoylsaxitoxin (dcSTX) were determined by liquid chromatography with fluorescence detection. Shellfish tissue was extracted with 30 mM HAc under ultrasonic, and cleanup of the extract was accomplished with a C18 solid-phase extraction column. Fluorescence oxidation of STX and dcSTX was carried out using 2% alkaline H2O2 solution. Chromatographic separation was performed on a C18 column (4.6 mm×250 mm, 5 μm) with elution of acetonitrile/0.1 M ammonium formate solution (5:95, v/v, pH 6) at a flow rate of 1.0 ml min−1. STX and dcSTX derivatives were completely separated in 7 min. The matrix-matched calibration graphs for STX and dcSTX were prepared by injecting the extracted blank spiked with increasing amounts of standards ranging from 0.01 to 2.0 μg g−1, giving an acceptable linearity (r = 0.9983 and r = 0.9989) over the test range. Average recoveries and relative standard deviations, by analyzing samples spiked at a level of 0.1, 0.8, and 1.6 μmg g−1, approximately ranged from 87% to 97% and from 8% to 13%, respectively. The detection limits (S/N=3) were 1.0 ng g−1 for STX and 0.3 ng g−1 for dcSTX. The proposed method was very sensitive, simple, fast, and reproducible. In addition, the structures of STX and dcSTX derivatives were elucidated by the quadrupole time-of-flight mass spectrometry.