Department of Pharmaceutical Sciences, College of Pharmacy, University of Illinois at Chicago, 833 South Wood Street, Chicago, IL 60612, USA; Pharmacognosy Institute, College of Pharmacy, University of Illinois at Chicago, 833 South Wood Street, Chicago, IL 60612, USA.
NMR Solutions Limited, Tulliportinkatu 3 A 8, 70100 Kuopio, Finland.
J Pharm Biomed Anal. 2022 May 30;214:114709. doi: 10.1016/j.jpba.2022.114709. Epub 2022 Mar 5.
The ICH guidelines recommend reporting thresholds for regular impurities in drug substances at the level of 0.05% or 0.03% (w/w) depending on the maximum daily intake. Therefore, any instrumental method of analysis applicable to the impurity analysis should be able to detect and quantify the analytes at those levels. This investigation was designed to verify the suitability of H NMR spectroscopy for the detection of impurities, as a first step in the process before attempting quantification. In order to minimize demand on equipment, this study employed a 400 MHz instrument for structural confirmation and signal assignments of choline (1) and O-(2-hydroxyethyl)choline (2), a known impurity. The limit of detection (LOD) of 2 in 10 mg of 1 was established as 0.01% on a 400 MHz instrument and 2% on a 60 MHz (benchtop) NMR spectrometer. Thus, impurities for which quantification is required are readily detected at 400 MHz or above. These results are in contrast to the widespread belief that H NMR sensitivity is insufficient for pharmaceutical impurity analysis. The choice of solvent was recognized as a critical parameter for H NMR LOD analysis. Furthermore, publicly available NMR raw data (HMDB) proved to be valuable for unveiling the otherwise cryptic information hidden in complex signal patterns via H NMR iterative Full Spin Analysis. Finally, the study uncovered the less noticed, yet characteristic, N-H coupling in the -N(CH) groups, adding strong arguments for the Raw NMR Data Initiative. Collectively, the data prove that the analytical capabilities of high-field NMR easily fulfill the ICH requirements for detection of impurity in the presence of an actual substance of interest which makes it a step closer to achieving regulatory standards.
ICH 指南建议根据每日最大摄入量，以 0.05%或 0.03%（w/w）的水平报告药物物质中常规杂质的报告阈值。因此，任何适用于杂质分析的仪器分析方法都应该能够在这些水平上检测和定量分析物。这项研究旨在验证 H NMR 光谱法用于检测杂质的适用性，作为在尝试定量之前的过程中的第一步。为了最大限度地减少对设备的需求，本研究采用 400 MHz 仪器对胆碱（1）和 O-（2-羟乙基）胆碱（2）（一种已知杂质）进行结构确认和信号分配。在 400 MHz 仪器上，1 中 2 的检测限（LOD）为 10mg 中的 0.01%，在 60 MHz（台式）NMR 光谱仪上为 2%。因此，需要定量的杂质可以在 400 MHz 或更高频率下轻松检测到。这些结果与广泛认为 H NMR 灵敏度不足以进行药物杂质分析的观点形成对比。溶剂的选择被认为是 H NMR LOD 分析的关键参数。此外，通过 H NMR 迭代全自旋分析，公开可用的 NMR 原始数据（HMDB）证明对于揭示复杂信号模式中隐藏的复杂信号模式中的隐藏信息非常有价值。最后，该研究揭示了-N(CH)基团中不太引人注意但特征性的 N-H 偶合，为原始 NMR 数据倡议增添了有力论据。总的来说，这些数据证明了高场 NMR 的分析能力很容易满足 ICH 对在实际感兴趣物质存在下检测杂质的要求，这使其更接近实现监管标准。
