Department of Applied Biotechnology and Food Science, Budapest University of Technology and Economics, Szent Gellért tér 4., H-1111, Budapest, Hungary.
Department of Biotechnology, Gedeon Richter Plc. Gyömrői út 19-21, H-1103, Budapest, Hungary.
J Pharm Biomed Anal. 2020 Feb 20;180:113060. doi: 10.1016/j.jpba.2019.113060. Epub 2019 Dec 19.
Chromatographic media play a crucial role in the downstream processing of biotechnology products. The physical and chemical properties of these processing aids are mostly monitored by expensive and time-consuming preparative tests, but spectroscopic techniques may also be used to measure chromatographic media samples. In this study, chromatographic media formulations used in downstream processes were investigated using attenuated total reflection Fourier transform infrared (ATR FT-IR) spectroscopy. Samples were measured both in original suspension form and after drying to examine the possibilities of a potential spectroscopic method without sample preparation. Principal component analysis (PCA) was employed to identify the spectral differences among the formulations with distinct support matrices and functional groups and soft independent modeling of class analogy (SIMCA) was performed to creating classification models for identification of chromatography media. To increase the number of samples in the SIMCA, simulated spectra were generated based on the experimental spectra. PCA models indicated that spectra of samples in original suspension form and after drying contained similar information about the chemical properties of chromatographic media samples. Moreover, during the classification of spectra based on SIMCA, both measurement techniques gave high sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) results. These results show that ATR FT-IR could be applied as a simple alternative method for monitoring the chromatography media samples. This technique is also feasible without sample preparation. Thereby the multi-hours drying steps may be omitted, the measurements can be performed in a few minutes, and the potential effects of sample preparations can be eliminated.
色谱介质在生物技术产品的下游处理中起着至关重要的作用。这些加工助剂的物理和化学性质主要通过昂贵且耗时的制备测试来监测,但也可以使用光谱技术来测量色谱介质样品。在这项研究中,使用衰减全反射傅里叶变换红外(ATR FT-IR)光谱法研究了下游工艺中使用的色谱介质配方。对原始悬浮液形式和干燥后的样品进行了测量,以检查无需样品制备的潜在光谱方法的可能性。主成分分析(PCA)用于识别具有不同支撑基质和官能团的配方之间的光谱差异,并进行软独立建模分类分析(SIMCA)以创建用于鉴定色谱介质的分类模型。为了增加 SIMCA 中的样本数量,根据实验光谱生成了模拟光谱。PCA 模型表明,原始悬浮液形式和干燥后的样品光谱均包含有关色谱介质样品化学性质的相似信息。此外,在基于 SIMCA 的光谱分类过程中,两种测量技术均给出了高灵敏度、特异性、阳性预测值(PPV)和阴性预测值(NPV)结果。这些结果表明,ATR FT-IR 可作为监测色谱介质样品的简单替代方法。该技术也无需样品制备即可实现。这样可以省略耗时数小时的干燥步骤,仅需几分钟即可完成测量,并且可以消除样品制备的潜在影响。