Extracellular carbonic anhydrase concentration at the sea-surface microlayer and 1 m depth during METEOR cruise M117, Baltic Sea

We developed an effective fluorometric technique to quantify extracellular carbonic anhydrase (eCA) present in natural seawater samples. The technique includes the separation of eCA from cells to achieve low detection limits through high signal : noise ratios. eCA was efficiently extracted from cell membranes by treatment with 0.1 M phosphate buffer containing 2.5 M NaCl. The free eCA specifically forms a fluorescent complex with dansylamide, and the detection limit of the complex is below 0.1 nM. We applied the technique to samples from different culture solutions and natural seawater collected from the Baltic Sea. We observed eCA concentrations to be in the range of 0.10-0.67 nM in natural seawater. The data indicated that this technique is very sensitive, accurate, and feasible for routine and shipboard measurement of eCA from natural seawater. It is therefore an effective and rapid tool to investigate the carbon acquisition of phytoplankton both in mono culture as well natural communities.

本研究开发了一种可定量天然海水样品中胞外碳酸酐酶（extracellular carbonic anhydrase, eCA）的高效荧光检测技术。该技术通过分离eCA与细胞以提升信噪比，从而实现低检出限。通过使用含2.5 mol/L氯化钠的0.1 mol/L磷酸盐缓冲液处理，可从细胞膜上高效提取eCA。游离态eCA可与丹磺酰胺（dansylamide）特异性结合形成荧光复合物，该复合物的检出限低于0.1 nM。本研究将该技术应用于不同培养液样品以及采自波罗的海的天然海水样品，测得天然海水样品中的eCA浓度范围为0.10~0.67 nM。实验数据表明，该技术灵敏度高、准确性强，可用于天然海水样品中eCA的常规检测与船载现场测定。因此，该技术可作为一种高效快速的工具，用于探究纯培养及自然群落中浮游植物的碳获取机制。