title: Development of a fast and selective method for the sensitive determination of anatoxin-a in lake waters using liquid chromatography-tandem mass spectrometry and phenylalanine-d5 as internal standard.
abstract: Anatoxin-a is a potent alkaloid neurotoxin produced by a number of cyanobacterial species and released in freshwaters during cyanobacterial blooms. Its high toxicity is responsible for several incidents of lethal intoxications of birds and mammals around the world; therefore anatoxin-a has to be regarded as a health risk and its concentration in lakes and water reservoirs should be monitored. Phenylalanine is a natural amino acid, also present in freshwaters, isobaric to anatoxin-a, with a very similar fragmentation pattern and LC retention. Since misidentification of phenylalanine as anatoxin-a has been reported in forensic investigations, special care must be taken in order to selectively determine traces of anatoxin-a in the presence of naturally occurring phenylalanine. A fast LC tandem MS method was developed by using a 1.8 microm 50 x 2.1 mm C18 column for the separation of anatoxin-a and phenylalanine, achieving a 3-min analysis time. Isotopically labelled phenylalanine-d(5) was employed as internal standard to compensate for electrospray ion suppression and sample preconcentration losses. Both compounds were preconcentrated 1,000-fold on a porous graphitic carbon solid-phase extraction (SPE) cartridge after adjustment of sample pH to 10.5. The method was validated by using lake water spiked at four different levels from 0.01 to 1 microg L(-1). Anatoxin-a recovery ranged from 73 to 97%, intra-day precision (RSD%) ranged from 4.2 to 5.9, while inter-day precision (RSD%) ranged from 4.2 to 9.1%. Limits of detection and quantification were 0.65 and 1.96 ng L(-1) respectively. The method was successfully applied for the detection of anatoxin-a in Greek lakes at concentrations ranging from less than 0.6 to 9.1 ng L(-1).
authors.affiliation: Laboratory of Analytical Chemistry, Department of Chemistry, University of Athens, Panepistimioupolis Zografou, 157 71 Athens, Greece.
authors.forename: Ioannis K
authors.initials: IK
authors.lastname: Dimitrakopoulos
authors.forename: Triantafyllos S
authors.initials: TS
authors.lastname: Kaloudis
authors.forename: Anastasia E
authors.initials: AE
authors.lastname: Hiskia
authors.forename: Nikolaos S
authors.initials: NS
authors.lastname: Thomaidis
authors.forename: Michael A
authors.initials: MA
authors.lastname: Koupparis
mesh_terms: D001427:Bacterial Toxins / Q000032:analysis; D002853:Chromatography, Liquid / Q000379:methods / Q000592:standards; D000087522:Cyanobacteria Toxins; D005618:Fresh Water / Q000032:analysis*; D006115:Greece; D057230:Limit of Detection; D008387:Marine Toxins; D008722:Methods; D010649:Phenylalanine; D015203:Reproducibility of Results; D052616:Solid Phase Extraction / Q000379:methods; D053719:Tandem Mass Spectrometry / Q000379:methods / Q000592:standards; D014326:Tropanes / Q000032:analysis*
chemical_list: D001427:Bacterial Toxins; D000087522:Cyanobacteria Toxins; D008387:Marine Toxins; D014326:Tropanes; D010649:Phenylalanine; C509783:anatoxin a