Use of high performance liquid chromatography to measure tetrodotoxin in serum and urine of poisoned patients.

Tetrodotoxin (TTX) poisoning is an infrequent but important problem in South-eastern Asia. Despite it being a considerable public health issue in some countries and its potential lethality there continues to be no readily available method for measuring TTX in urine or serum. Previously published methods have used immunoaffinity chromatography, or the conversion of TTX to its C9-base derivative for measurement by mass spectrometry. A simple and reproducible method was developed using solid phase extraction cartridges to clean up serum and urine samples from TTX-poisoned patients, and the subsequent analysis of the samples by high performance liquid chromatography with post-column derivatisation and fluorescence detection. Minimum quantifiable concentrations of TTX were 5 and 20 ng/ml for serum and urine, respectively. Precision and accuracy of the assay were 13 and 15%, respectively. The standard curves were linear in the range of 20-300 ng/ml for urine and 5-20 ng/ml for serum. TTX was quantified in six samples of urine and six samples of serum from seven patients who ingested common toadfish and had clinical effects consistent with TTX poisoning. TTX was detected in all urine samples but in only one serum sample. Using this method confirmation of TTX poisoning will be far simpler and readily available. A 24 h urine collection in the period immediately following poisoning is likely to be the most sensitive test for TTX poisoning. With appropriately collected and timed serum and urine specimens it will be possible to properly define the pharmacokinetics of TTX in humans.