Doak DL, Phillips JA
Department of Chemical Engineering, Lehigh University, Bethlehem, Pennsylvania 18015, USA.
Biotechnol Prog. 1999 May;15(3):529-39. doi: 10.1021/bp990039b.
A diamond composition ATR probe was used in situ to obtain IR spectra on replicate Escherichia coli fermentations involving a complex medium. The probe showed excellent stability over a 6-month operating period and was unaffected by either agitation or aeration. The formation of an unknown was observed from IR spectra obtained during the sterilization; subsequent experiments proved this to be a reaction product between yeast extract and the phosphates used as buffer salts. Partial-least-squares-based calibration/prediction models were developed for both glucose and acetate using in-process samples. The resulting models had prediction errors of +/-0.26 and +/-0.75 g/L for glucose and acetic acid, respectively, errors which were statistically equivalent to the estimated experimental errors in the reference measurements. Relative concentration profiles for the unknown formed during sterilization could be generated either by tracking peak height at an independent wavelength or by self-modeling curve resolution of the spectral region overlapping that of glucose. These profiles indicated that this compound was metabolized simultaneously with glucose; upon depletion of the glucose, when the microorganism switched to consumption of acetic acid, utilization continued but at a lower rate. The data presented provide an extensive characterization of the performance characteristics of this in situ analysis and clearly demonstrate its utility not just in the quantitative measurement of multiple known species but in the qualitative evaluation of unknown species.
使用金刚石成分的衰减全反射（ATR）探头原位获取涉及复杂培养基的重复大肠杆菌发酵过程的红外光谱。该探头在6个月的运行期内表现出出色的稳定性，不受搅拌或通气的影响。在灭菌过程中获得的红外光谱中观察到一种未知物的形成；后续实验证明这是酵母提取物与用作缓冲盐的磷酸盐之间的反应产物。使用过程中的样品为葡萄糖和乙酸开发了基于偏最小二乘法的校准/预测模型。所得模型对葡萄糖和乙酸的预测误差分别为±0.26和±0.75 g/L，这些误差在统计学上与参考测量中的估计实验误差相当。灭菌过程中形成的未知物的相对浓度曲线可以通过跟踪独立波长处的峰高或通过对与葡萄糖重叠的光谱区域进行自建模曲线分辨来生成。这些曲线表明该化合物与葡萄糖同时被代谢；当葡萄糖耗尽时，微生物转向消耗乙酸，其利用仍在继续，但速率较低。所呈现的数据对这种原位分析的性能特征进行了广泛的表征，并清楚地证明了其不仅在多种已知物质的定量测量中有用，而且在未知物质的定性评估中也有用。
