Abstract
The aim of this study was to develop a practical method to analyze tetrodotoxin (TTX), quantitatively, from postmortem specimens, not only blood and urine, but also organs. Extraction was achieved with 2% acetic acid and the use of an anion-exchange solid-phase extraction (SPE) cartridge. The quantitation method was a standard addition method with a calibration curve consisting of at least 3 points and an internal standard, voglibose (VOG). Separation by LC-MS/MS was achieved using a Luna HILIC (Phenomenex) column. The mobile phase was acetonitrile: 5 mM ammonium formate buffer (95:5), delivered at 0.2 mL/min. The selected reaction monitoring (SRM) transitions for TTX and VOG were m/z 320>302 and m/z 268>92, respectively. Cleaner extracts were achieved by using a lipid removal cartridge and washing with heptane. The addition of steps to remove interfering components that are prominent in postmortem samples aided in successful analysis. The HILIC column improved the retention of TTX to greater than 2 min to avoid the area where ion suppression has its greatest effect. Also, the use of anion-exchange SPE lessened the influence of acetic acid used during extraction. By using this method, we were able to quantitate low levels of TTX in postmortem specimens.
