Sensitive determination of tetrodotoxin in human plasma by gas chromatography/mass spectrometry

A sensitive and reliable method for determination of tetrodotoxin (TTX) in human plasma by gas chromatography/ mass spectrometry (GC/MS) was developed. TTX was extracted from plasma with 2% acetic acid in a mixture of methanol and acetonitrile (8:2), passed through the first cartridge, and then transformed into 2-amino-6-hydroxymethyl-8-hydroxyquinazoline (C₉-base) with 1 M potassium hydroxide. The C₉-base was purified with the second cartridge, converted to the TMS derivative together with β-estradiol (IS) and then applied to a GC/MS instrument. Three kinds of cartridges, Sep-Pak C₁₈, Sep-Pak tC₁₈ and Sep-Pak PS-2, were all found to be equally useful when used as the first cartridge, but the Sep-Pak PS-2 gave the best recovery when used as the second cartridge. The method described here enabled the quantitation of TTX in human plasma in the range of 0.5-100 ng/ml with the lowest limit of detection of 0.5 ng/ml.