Chemometric evaluation of on-line high-pressure liquid chromatography in mammalian cell cultures: Analysis of amino acids and glucose
Chemometric evaluation of on-line high-pressure liquid chromatography in mammalian cell cultures: Analysis of amino acids and glucose
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DOI：
10.1002/bit.10116
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发表时间：
2002-03-05
影响因子：
3.8
通讯作者：
Cacia, J
中科院分区：
文献类型：
--
作者：
Larson, TM;Gawlitzek, M;Cacia, J
建立了一种能够在线检测氨基酸和碳水化合物的高压液相色谱系统，用于研究哺乳动物细胞培养系统中的代谢。高效液相色谱法采用阴离子交换层析和积分脉冲安培检测法。该方法能同时测定19种氨基酸和葡萄糖，完整的测定时间为65min。在实际细胞培养中，该方法被证明对监测17种氨基酸和葡萄糖是有用的。没有准确监测的两种氨基酸是精氨酸和赖氨酸，可能是因为它们在柱的空隙体积附近洗脱。该系统被用来研究不同细胞培养过程中代谢的变异性，以及葡萄糖和谷氨酰胺限制对单个细胞培养过程的影响。化学计量学分析被用来从这些实验产生的高度相关的多变量数据集中得出有统计学意义的结论。利用化学计量学，不同过程之间的差异与七种氨基酸摄取率的差异有关。同样，乳酸浓度、细胞密度和天冬氨酸摄取率与葡萄糖和谷氨酰胺限制有关。还考虑了营养限制对谷氨酸、丙氨酸和铵的影响。(C)2002年John Wiley Sons，Inc.
An on-line high-pressure liquid chromatography (HPLC) system capable of measuring amino acids and carbohydrates was used to study metabolism in mammalian cell culture systems. The HPLC method utilized anion-exchange chromatography followed by integrated pulsed amperometric detection. The method is capable of measuring 19 amino acids plus glucose with a complete method time of 65 min. In actual cell cultures, the method was shown to be useful for monitoring 17 amino acids plus glucose. The two amino acids that were not accurately monitored were arginine and lysine, possibly due to their elution near the void volume of the column. The HPLC system was used to study variability in metabolism across different cell culture processes, as well as the effect of glucose and glutamine limitation on a single cell culture process. Chemometric analysis was used to draw statistically meaningful conclusions from the highly correlated, multivariate data set that resulted from these experiments. Using chemometrics, variation between processes was linked to differences in uptake rates of seven amino acids. Similarly, lactate concentration, cell density, and aspartate uptake rate were linked to glucose and glutamine limitation. The effect of nutrient limitation on glutamate, alanine, and ammonium was also considered. (C) 2002 John Wiley Sons, Inc.