Verification and quantification of saxitoxin from algal samples using fast and validated hydrophilic interaction liquid chromatography–tandem mass spectrometry method

Abstract

Hydrophilic interaction liquid chromatography–tandem mass spectrometry (HILIC–MS/MS) method was validated with algal samples for verification and quantification of saxitoxin (STX), a potent neurotoxin which is listed in the Chemical Weapons Convention (CWC) in Schedule 1A. Isocratic elution, conventional bore HILIC column and high flow rate together with accurate post-column splitter provided detection of STX in 6.5min with total analysis time of 9min per sample. STX analogue, gonyautoxin 1 (GTX 1) was used as an internal standard. Sample preparation of freeze-dried algae included liquid extraction and centrifugal filtering with mean recovery of 99.9% at concentration level of 10ng/ml (n=3). Retention times for STX and GTX 1 were 6.47±0.03min and 4.44±0.01min (n=45), respectively. Four diagnostic product ions were used for reliable verification of saxitoxin. Method was found to be precise and linear (R2=0.9714 and R2=0.9768) in concentration ranges of 5–50ng/ml and 25–200ng/ml, respectively. For saxitoxin, calculated LOD was 3ng/ml and LLOQ 11ng/ml. Validation was conducted using spiked algal matrix since this method is not only needed for verification analysis for the CWC but also for safety analysis of other environmental samples for presence of STX. Identification criteria for verification of STX with HILIC–MS/MS method are discussed.