Detection and quantification of pufferfish tetrodotoxin by high-performance liquid chromatography

Abstract

Tetrodotoxin (TTX) is a neurotoxin mainly found in pufferfish. Although food poisoning caused by TTX has been reported in recent years, TTX measurement in biological samples derived from patients has rarely been performed at medical facilities. Therefore, we attempted to establish a high-performance liquid chromatography (HPLC) analytical method using an anion-exchange column to detect and quantify TTX. First, TTX samples (0.5–50 μg/mL) diluted in citrate buffer solution were analyzed by HPLC. According to this method, TTX chromatograms were detected in the retention time range of 0.1–0.4 min. Furthermore, TTX in citrate buffer could be detected in the concentration range of 1.0–50 μg/mL, and the standard curve was successfully created on the basis of the obtained results. Next, we examined whether TTX diluted in human serum or urine could be detected and quantified. Despite deproteinization, TTX in serum could not be measured. In urine samples, TTX was detected in the concentration range of 5.0–50 μg/mL; however, the calculated value was lower than the actual TTX concentration. Next, the calculated TTX concentration was corrected by applying the recreated standard curve based on the measurement results of TTX samples diluted in urine. In conclusion, this HPLC analytical method may be adapted to urine samples from patients with TTX poisoning.