Furey Ambrose, Crowley Janet, Lehane Mary, James Kevin J
PROTEOBIO, Mass Spectrometry Centre for Proteomics and Biotoxin Research, Department of Chemistry, Cork Institute of Technology, Bishopstown, Cork, Ireland.
Rapid Commun Mass Spectrom. 2003;17(6):583-8. doi: 10.1002/rcm.932.
Anatoxin-a (AN) and homoanatoxin-a (HMAN) are potent neurotoxins produced by a number of cyanobacterial species. A new, sensitive liquid chromatography/multiple tandem mass spectrometry (LC/MS(n)) method has been developed for the determination of these neurotoxins. The LC system was coupled, via an electrospray ionisation (ESI) source, to an ion-trap mass spectrometer in positive ion mode. The M+H ions at m/z 166 (anatoxin-a) and m/z 180 (homoanatoxin-a) were used as the precursor ions for multiple MS experiments. MS(2)bond;MS(4) spectra displayed major fragment ions at m/z 149 (AN), 163 (HMAN), assigned to Mbond;NH(3)+H; m/z 131 (AN), 145 (HMAN), assigned to Mbond;NH(3)bond;H(2)O+H, and m/z 91 C(7)H(7). Although the chromatographic separation of these neurotoxins is problematic, reversed-phase LC, using a C(18) Luna column, proved successful. Calibration data for anatoxin-a using spiked water samples (10 mL) in LC/MS(n) modes were: LC/MS (25-1000 microg/L), r(2) = 0.998; LC/MS(2) (5-1000(microg/L), r(2) = 0.9993; LC/MS(3) (2.5-1000 microg/L), r(2) = 0.9997. Reproducibility data (% RSD, N = 3) for each LC/MS(n) mode ranged between 2.0 at 500 microg/L and 7.0 at 10 microg/L. The detection limit (S/N = 3) for AN was better than 0.03 ng (on-column) for LC/MS(3) which corresponded to 0.6 microg/L.
类毒素-a（AN）和高类毒素-a（HMAN）是多种蓝藻物种产生的强效神经毒素。已开发出一种新的、灵敏的液相色谱/多重串联质谱（LC/MS(n)）方法来测定这些神经毒素。液相色谱系统通过电喷雾电离（ESI）源与处于正离子模式的离子阱质谱仪相连。质荷比为166（类毒素-a）和180（高类毒素-a）的M+H离子用作多重质谱实验的前体离子。MS(2)；MS(4)谱显示质荷比为149（AN）、163（HMAN）的主要碎片离子，归属于M-NH₃+H；质荷比为131（AN）、145（HMAN）的碎片离子，归属于M-NH₃-H₂O+H，以及质荷比为91的C₇H₇。尽管这些神经毒素的色谱分离存在问题，但使用C₁₈ Luna柱的反相液相色谱证明是成功的。在LC/MS(n)模式下，对添加了类毒素-a的水样（10 mL）的校准数据如下：LC/MS（25 - 1000 μg/L），r² = 0.998；LC/MS(2)（5 - 1000 μg/L），r² = 0.9993；LC/MS(3)（2.5 - 1000 μg/L），r² = 0.9997。每种LC/MS(n)模式的重现性数据（% RSD，N = 3）在500 μg/L时为2.0，在10 μg/L时为7.0。LC/MS(3)对AN的检测限（S/N = 3）优于0.03 ng（柱上），相当于0.6 μg/L。
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