Fischer W J, Garthwaite I, Miles C O, Ross K M, Aggen J B, Chamberlin A R, Towers N R, Dietrich D R
Nestlé Research Center, Nestec Ltd., Vers-chez-les-Blanc, Lausanne, Switzerland.
Environ Sci Technol. 2001 Dec 15;35(24):4849-56. doi: 10.1021/es011182f.
Cyanobacteria (blue-green algae) (e.g., Microcystis and Nodularia spp.) capable of producing toxic peptides are found in fresh and brackish water worldwide. These toxins include the microcystin (MC) heptapeptides (>60 congeners) and the nodularin pentapeptides (ca. 5 congeners). Cyanobacterial cyclic peptide toxins are harmful to man, other mammals, birds, and fish. Acute exposure to high concentrations of these toxins causes liver damage, while subchronic or chronic exposure may promote liver tumor formation. The detection of cyclic peptide cyanobacterial toxins in surface and drinking waters has been hampered by the low limits of detection required and that the present routine detection is restricted to a few of the congeners. The unusual beta-amino acid ADDA (4E,6E-3-amino-9-methoxy-2,6,8-trimethyl-10-phenyldeca-4,6-dienoic acid) is present in most (>80%) of the known toxic penta- and heptapeptide toxin congeners. Here, we report the synthesis of two ADDA-haptens, the raising of antibodies to ADDA, and the development of a competitive indirect ELISA for the detection of microcystins and nodularins utilizing these antibodies. The assay has a limit of quantitation of 0.02-0.07 ng/mL (depending on which congeners are present), lower than the WHO-proposed guideline (1 ng/mL) for drinking water, irrespective of the sample matrix (raw water, drinking water, or pure toxin in PBS). This new ELISA is robust, can be performed without sample preconcentration, detects toxins in freshwater samples at lower concentrations than does the protein phosphatase inhibition assay, and shows very good cross-reactivity with all cyanobacterial cyclic peptide toxin congeners tested to date (MC-LR, -RR, -YR, -LW, -LF, 3-desmethyl-MC-LR, 3-desmethyl-MC-RR, and nodularin).
能产生有毒肽的蓝藻（蓝绿藻）（如微囊藻属和节球藻属）在世界各地的淡水和微咸水中均有发现。这些毒素包括微囊藻毒素（MC）七肽（>60种同系物）和节球藻毒素五肽（约5种同系物）。蓝藻环肽毒素对人类、其他哺乳动物、鸟类和鱼类有害。急性接触高浓度这些毒素会导致肝脏损伤，而亚慢性或慢性接触可能会促进肝脏肿瘤形成。地表水和饮用水中环肽蓝藻毒素的检测一直受到所需检测限低以及目前常规检测仅限于少数同系物的阻碍。大多数（>80%）已知的有毒五肽和七肽毒素同系物中都存在不寻常的β-氨基酸ADDA（4E,6E-3-氨基-9-甲氧基-2,6,8-三甲基-10-苯基癸-4,6-二烯酸）。在此，我们报告了两种ADDA半抗原的合成、针对ADDA抗体的产生以及利用这些抗体开发用于检测微囊藻毒素和节球藻毒素的竞争性间接酶联免疫吸附测定。该测定的定量限为0.02 - 0.07 ng/mL（取决于存在哪些同系物），低于世界卫生组织提议的饮用水指导值（1 ng/mL），无论样品基质是原水、饮用水还是PBS中的纯毒素。这种新的酶联免疫吸附测定性能稳定，无需样品预浓缩即可进行，能检测淡水样品中浓度低于蛋白质磷酸酶抑制测定法的毒素，并且与迄今测试的所有蓝藻环肽毒素同系物（MC-LR、-RR、-YR、-LW、-LF、3-去甲基-MC-LR、3-去甲基-MC-RR和节球藻毒素）都具有很好的交叉反应性。
