To better understand pH-dependence of endogenous fluorescence of algae, we employed spectroscopy and microscopy methods, including advanced time-resolved fluorescence imaging microscopy (FLIM), using green algae Chlorella sp. as a model system. Absorption spectra confirmed two peaks, at 400-420 nm and 670 nm. Emission was maximal at 680 nm, with smaller peaks between 520 and 540 nm. Acidification led to a gradual decrease in the red fluorescence intensity with the maximum at 680 nm when excited by 450 nm laser. FLIM measurements, performed using 475 nm picoseconds excitation, uncovered that this effect is accompanied by a shortening of the tau1 fluorescence lifetime. Under severe acidification, we also noted an increase in the green fluorescence with a maximum between 520-540 nm and a shift toward 690-700 nm of the red fluorescence, accompanied by prolongation of the tau2 fluorescence lifetime. Gathered data increase our knowledge on the responsiveness of algae to acidification and indicate that endogenous fluorescence derived from chlorophylls can potentially serve as a biosensing tool for monitoring pH change in its natural environment.

中文翻译：

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时间分辨的内源叶绿素荧光对pH的敏感性：小球藻的研究。藻类。

为了更好地了解藻类内源性荧光的pH依赖性，我们采用了光谱学和显微镜方法，包括使用绿藻小球藻属的先进的时间分辨荧光成像显微镜（FLIM）。作为模型系统。吸收光谱证实在400-420nm和670nm处有两个峰。发射在680 nm处最大，在520和540 nm之间有较小的峰。酸化导致红色荧光强度逐渐降低，当被450 nm激光激发时，最大荧光强度在680 nm处降低。使用475 nm皮秒激发进行的FLIM测量发现，这种效应伴随着tau1荧光寿命的缩短。在严重的酸化条件下，我们还注意到绿色荧光增加，最大值在520-540 nm之间，红色荧光向690-700 nm转移，伴随着tau2荧光寿命的延长。收集的数据增加了我们对藻类对酸化反应性的了解，并表明源自叶绿素的内源性荧光可潜在地用作监测其自然环境中pH变化的生物传感工具。