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Evaluation of fluorimetric pH sensors for bioprocess monitoring at low pH

机译：评估低pH下用于生物过程监测的荧光pH传感器

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Optical chemical sensors are the standard for pH monitoring in small-scale bioreactors such as microtiter plates, shaking flasks or other single-use bioreactors. The dynamic pH range of the so far commercially available fluorescent pH sensors applied in small-scale bioreactors is restricted to pH monitoring around neutral pH, although many fermentation processes are performed at pH < 6 on industrial scale. Thus, two new prototype acidic fluorescence pH sensors immobilized in single-use stirred-tank bioreactors, one with excitation at 470 nm and emission at 550 nm (sensor 470/550) and the other with excitation at 505 nm and emission at 600 nm (sensor 505/600), were characterized with respect to dynamic ranges and operational stability in representative fermentation media. Best resolution and dynamic range was observed with pH sensor 505/600 in mineral medium (dynamic range of 3.9 < pH < 7.2). Applying the same pH sensors to complex medium results in a drastic reduction of resolution and dynamic ranges. Yeast extract in complex medium was found to cause background fluorescence at the sensors' operating wavelength combinations. Optical isolation of the sensor by adding a black colored polymer layer above the sensor spot and fixing an aperture made of adhesive photoresistant foil between the fluorescence reader and the transparent bottom of the polystyrene reactors enabled full re-establishment of the sensor's characteristics. Reliability and operational stability of sensor 505/600 was shown by online pH monitoring (4.5 < pH < 5.8) of parallel anaerobic batch fermentations of Clostridium acetobutylicum for the production of acetone, butanol and ethanol (ABE) with offline pH measurements with a standard glass electrode as reference.
机译：光学化学传感器是在小型生物反应器（例如微量滴定板，摇瓶或其他一次性生物反应器）中进行pH监测的标准。尽管许多发酵过程是在工业规模的pH <6下进行的，但迄今为止，在小型生物反应器中使用的市售荧光pH传感器的动态pH范围仅限于中性pH左右的pH监测。因此，将两个新的原型酸性荧光pH传感器固定在一次性搅拌罐式生物反应器中，一个在470 nm激发并在550 nm处发射（传感器470/550），另一个在505 nm激发并在600 nm处发射（关于代表性发酵培养基中的动态范围和操作稳定性，对传感器505/600进行表征。使用pH传感器505/600在矿物介质中观察到最佳分辨率和动态范围（动态范围3.9 H <7.2）。将相同的pH传感器应用于复杂的介质会大大降低分辨率和动态范围。发现在复杂培养基中的酵母提取物会在传感器的工作波长组合下引起背景荧光。通过在传感器光斑上方添加黑色聚合物层并在荧光读取器和聚苯乙烯反应器的透明底部之间固定一个由粘性耐光箔制成的孔来实现传感器的光学隔离，从而可以完全恢复传感器的特性。通过对丙酮丁醇梭菌生产的丙酮，丁醇和乙醇（ABE）进行平行厌氧分批发酵的在线pH监测（4.5 H <5.8）在线pH监测（4.5 H <5.8）来显示传感器505/600的可靠性和操作稳定性，并使用标准玻璃进行离线pH测量电极作为参考。

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