Ross Keri Lyn, Tu Travis T, Smith Sean, Dalluge Joseph J
Cargill Global Food Technology Group, Cargill Incorporated, P.O. Box 5702, Minneapolis, Minnesota 55440-5702, USA.
Anal Chem. 2007 Jul 1;79(13):4840-4. doi: 10.1021/ac0624243. Epub 2007 May 26.
A method has been developed for rapid quantification of organic acids using ultraperformance liquid chromatography/electrospray-tandem mass spectrometry (UPLC/ESI-MS-MS) to monitor the metabolism of 10 organic acids during microbial fermentation. Because comprehensive chromatographic separation is not required, analysis time is less than traditional ion chromatography assays, with complete organic acid analyses by UPLC/ESI-MS-MS being achieved in less than 3 min. Quantification is accomplished using nine isotopically labeled organic acids as internal standards. Intrasample precisions for organic acid measurements in fermentation supernatants using this method average 8.9% (RSD). Calibration curves are linear over the range of 0.06-100 microg/mL, and detection limits are estimated at 0.06-1 microg/mL. This method has the potential to demonstrate correlation of organic acid consumption and production by microorganisms with observed growth profiles, novel media formulations, and cellular growth events. Data visualization software has been used to profile organic acid levels during fermentation and correlate these profiles to nutrient supplementation protocols employed during microbial production. The potential use of this capability in computational modeling and simulation of microbial metabolism to accelerate the bioprocess development cycle is recognized.
已开发出一种使用超高效液相色谱/电喷雾串联质谱法（UPLC/ESI-MS-MS）快速定量有机酸的方法，以监测微生物发酵过程中10种有机酸的代谢情况。由于不需要全面的色谱分离，分析时间比传统离子色谱分析短，UPLC/ESI-MS-MS在不到3分钟内即可完成完整的有机酸分析。使用9种同位素标记的有机酸作为内标进行定量。使用该方法测量发酵上清液中有机酸的样品内精密度平均为8.9%（相对标准偏差）。校准曲线在0.06 - 100μg/mL范围内呈线性，检测限估计为0.06 - 1μg/mL。该方法有可能证明微生物消耗和产生的有机酸与观察到的生长曲线、新型培养基配方和细胞生长事件之间的相关性。数据可视化软件已用于描绘发酵过程中的有机酸水平，并将这些曲线与微生物生产过程中采用的营养补充方案相关联。人们认识到这种能力在微生物代谢的计算建模和模拟中潜在的用途，可加速生物工艺开发周期。
