Chlorophyll-a and phycocyanin concentrations from a Nodularia spumigena bloom event on Lake Bante in Wilhelmshaven, Germany

Lake Bant in Germany was surveyed on 25 August 2022 and the gathered water samples were filtered through 0.2 µm pore sized Whatman Nuclepore Track-Etch membranes with a diameter of 47 mm. The filters were frozen and stored at -80 °C until further analyses. These frozen filters were transferred to a 20 mL glass vial followed by addition of 5 mL 1 × phosphate buffered saline with a pH = 7 to obtain pigments concentrations. An ultrasonic bath filled with crushed ice was used to sonicate the samples for 4 times in 60 second pulses with 60 second pauses in between the sonication. The sample glass vials were then centrifuged and 3 × 250 µL of the supernatant was transferred onto a 96 microwell plate. Sample absorbance was obtained using a BioTek Instruments Synergy H1 Hybrid Multi-mode microplate reader. Phycocyanin concentrations (µg/L) were derived following the recommended steps (Horváth et al., 2013). The remaining supernatant was decanted and the filters were refrozen at -22°C. Samples were freeze-dried for 24 h and then 5 mL of 99.5% ethanol were added to each glass vial. The alcoholic extracts were prepared as explained above. Again the samples were centrifuged and the supernatant from each sample was measured in the microplate reader. Chlorophyll a concentrations (mg/L) was estimated as described by Ritchie, 2008.

2022年8月25日对德国班特湖（Lake Bant）开展野外调研，采集的水样经孔径0.2 µm的Whatman Nuclepore径迹蚀刻膜（Track-Etch membranes，直径47 mm）过滤。滤膜经冷冻后保存于-80 ℃以待后续分析。将上述冷冻滤膜转移至20 mL玻璃样品瓶中，加入5 mL pH为7的1×磷酸盐缓冲液以测定色素浓度。使用装有碎冰的超声浴槽对样品进行超声处理：以60秒为一个超声脉冲，共进行4次脉冲，每次脉冲间隔60秒。随后将样品玻璃瓶离心，取3份250 µL的上清液转移至96孔微孔板（96 microwell plate）中。使用伯腾仪器（BioTek Instruments）Synergy H1混合型多功能微孔板读数仪测定样品吸光度。藻蓝蛋白浓度（单位：µg/L）按照Horváth等人2013年推荐的实验步骤计算得到。剩余上清液被倾弃，滤膜重新冷冻保存于-22 ℃。样品经24小时冷冻干燥后，向每个玻璃样品瓶中加入5 mL 99.5%乙醇。按照前述方法制备乙醇提取物，再次离心后取各样本上清液，使用微孔板读数仪进行检测。叶绿素a浓度（单位：mg/L）参照Ritchie 2008年的方法进行估算。