Polyphosphates have occurred in living cells early in evolution and microalgae contain these important polymers in their cells. Progress in research of polyphosphate metabolism of these ecologically as well as biotechnologically important microorganisms is hampered by the lack of rapid quantification methods. Experiments with the green alga Chlorella vulgaris presented here compared polyphosphate extraction in water, methanol-chloroform, and phenol-chloroform followed by polyphosphate purification by binding to silica columns or ethanol precipitation. The phenol-chloroform extraction of C. vulgaris followed by ethanol precipitation of polyphosphate was shown to be superior to the other tested method variants. Recovery test of added polyphosphate standard to algal biomass showed that the method is accurate. Using this biochemical assay as a validated reference, we show that 2-dimensional, confocal Raman microscopy can serve as a linear proxy for polyphosphate in C. vulgaris with R² up to 0.956. With this, polyphosphate quantification can be shortened by use of Raman microscopy from days to hours and, additionally, information about intracellular distribution of polyphosphate and heterogeneity among individual cells in algal culture can be obtained. This offers new insights into the dynamics and role of these polymers crucial for phosphorus uptake and storage. This analytical capability is of particular practical importance because algae aid phosphorus sequestration from wastewater and the thus enriched biomass may serve as organic fertilizer. Both these applications have a strong potential in a future sustainable, circular bioeconomy.

中文翻译：

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通过拉曼显微镜和参考酶法定量测定微藻中的多磷酸盐

聚磷酸盐已在进化早期的活细胞中发生，微藻类在其细胞中包含这些重要的聚合物。缺乏快速定量方法阻碍了这些在生态学和生物技术上重要的微生物的多磷酸盐代谢研究的进展。这里介绍的绿藻小球藻的实验比较了在水中，甲醇-氯仿和苯酚-氯仿中提取多磷酸盐，然后通过与硅胶柱结合或乙醇沉淀纯化多磷酸盐。寻常衣藻的苯酚-氯仿提取随后用乙醇沉淀多磷酸盐被证明优于其他测试方法。藻类生物质中添加多磷酸盐标准品的回收率测试表明该方法是准确的。使用这种生化测定作为验证的参考，我们表明，2维的，共焦拉曼显微镜检查可作为在多磷酸盐的线性代理普通小球藻用- [R ²高达0.956。这样，通过使用拉曼显微术可以将多磷酸盐的定量从几天缩短到几小时，此外，可以获得关于多磷酸盐的细胞内分布和藻类培养物中单个细胞之间的异质性的信息。这提供了对这些聚合物的动力学和作用的新见解，这些动力学和作用对于磷的吸收和储存至关重要。这种分析能力特别具有实际意义，因为藻类有助于从废水中隔离磷，因此富集的生物质可以用作有机肥料。这两种应用在未来的可持续循环生物经济中都具有强大的潜力。