Zamlynny Lydia, Morris Hannah M, Giddings Sabrina D, Kollatz Johannes, Niedermeyer Timo H J, Jamieson Rob C, Beach Daniel G
Department of Civil and Resource Engineering, Dalhousie University, 6299 South St, Halifax, NS, Canada.
Metrology Research Centre, National Research Council of Canada, 1411 Oxford St, Halifax, NS, B3H 3Z1, Canada.
Anal Bioanal Chem. 2025 Mar 27. doi: 10.1007/s00216-025-05829-9.
Cyanobacteria are prokaryotic organisms that can form large monospecific blooms, which pose a risk to human and animal health as some species produce toxic secondary metabolites called cyanotoxins. Multiclass cyanotoxin analysis is challenging due to varying chemical and physical properties between classes, as well as potentially large numbers of analogues within each class. Incorporating anatoxins (ATXs) into multiclass methods can be particularly challenging due to their small molecular size, potential interferences, polarity, and a lack of chemical standards for most analogues. Here, we present the development of a multiclass LC-MS/MS method and a quantitative calibration solution for aetokthonotoxin (AETX), an emerging cyanotoxin linked to mass mortalities of bald eagles in the Eastern United States. The developed method is capable of detecting 17 microcystins (MCs), nodularin-R, three cylindrospermopsins (CYNs), AETX, and 17 ATXs, including recently tentatively identified 10-hydroxy analogues. Analytes were identified by retention time and product ion ratio matching with available standards. The method was evaluated with respect to limits of detection (LODs), linear range, accuracy, and precision using neat and matrix matched standards. LODs in wet cyanobacterial biofilms ranged from 0.14 ng/g for CYN to 2.8 ng/g for [Dha]MC-LR with accuracies ranging from 65% for [Leu]MC-LY to 116% for CYN. Finally, the method's application was demonstrated through analysis of cyanobacterial field samples, a dietary supplement matrix reference material, and passive sampler extracts to assess versatility within different matrices.
蓝藻是原核生物，能够形成大规模的单种藻华，由于某些物种会产生称为蓝藻毒素的有毒次生代谢产物，这对人类和动物健康构成风险。多类蓝藻毒素分析具有挑战性，因为不同类别之间的化学和物理性质各不相同，而且每类中可能存在大量类似物。将类毒素（ATXs）纳入多类分析方法尤其具有挑战性，因为它们分子尺寸小、存在潜在干扰、具有极性，并且大多数类似物缺乏化学标准品。在此，我们展示了一种多类液相色谱-串联质谱（LC-MS/MS）方法的开发以及一种针对埃托毒素（AETX）的定量校准溶液，AETX是一种新出现的蓝藻毒素，与美国东部白头海雕的大量死亡有关。所开发的方法能够检测17种微囊藻毒素（MCs）、节球藻毒素-R、三种柱孢藻毒素（CYNs）、AETX以及17种ATXs，包括最近初步鉴定出的10-羟基类似物。通过保留时间和产物离子比率与现有标准品匹配来鉴定分析物。使用纯品和基质匹配标准品对该方法的检测限（LODs）、线性范围、准确度和精密度进行了评估。在湿蓝藻生物膜中的检测限范围为，CYN为0.14 ng/g，[Dha]MC-LR为2.8 ng/g，准确度范围为，[Leu]MC-LY为65%，CYN为116%。最后，通过对蓝藻现场样品、膳食补充剂基质参考物质和被动采样器提取物的分析，展示了该方法的应用，以评估其在不同基质中的通用性。
