Department of Biomedical & Pharmaceutical Sciences, Harry and Diane Rinker Health Science Campus, Chapman University, Irvine, CA, 92618, USA.
Department of Pharmacology and Toxicology, Faculty of Pharmacy, Northern Border University, Rafha, Kingdom of Saudi Arabia.
Sci Rep. 2021 Feb 1;11(1):2703. doi: 10.1038/s41598-021-81817-0.
During drug development, evaluation of drug and its metabolite is an essential process to understand drug activity, stability, toxicity and distribution. Liquid chromatography (LC) coupled with mass spectrometry (MS) has become the standard analytical tool for screening and identifying drug metabolites. Unlike LC/MS approach requiring liquifying the biological samples, we showed that spectral imaging (or spectral microscopy) could provide high-resolution images of doxorubicin (dox) and its metabolite doxorubicinol (dox'ol) in single living cells. Using this new method, we performed measurements without destroying the biological samples. We calculated the rate constant of dox translocating from extracellular moiety into the cell and the metabolism rate of dox to dox'ol in living cells. The translocation rate of dox into a single cell for spectral microscopy and LC/MS approaches was similar (~ 1.5 pM min cell). When compared to spectral microscopy, the metabolism rate of dox was underestimated for about every 500 cells using LC/MS. The microscopy approach further showed that dox and dox'ol translocated to the nucleus at different rates of 0.8 and 0.3 pM min, respectively. LC/MS is not a practical approach to determine drug translocation from cytosol to nucleus. Using various methods, we confirmed that when combined with a high-resolution imaging, spectral characteristics of a molecule could be used as a powerful approach to analyze drug metabolism. We propose that spectral microscopy is a new method to study drug localization, translocation, transformation and identification with a resolution at a single cell level, while LC/MS is more appropriate for drug screening at an organ or tissue level.
在药物开发过程中，评估药物及其代谢物是了解药物活性、稳定性、毒性和分布的必要过程。液相色谱（LC）与质谱（MS）联用已成为筛选和鉴定药物代谢物的标准分析工具。与需要将生物样本液化的 LC/MS 方法不同，我们表明，光谱成像（或光谱显微镜）可以提供单个活细胞中阿霉素（dox）及其代谢物阿霉素醇（dox'ol）的高分辨率图像。使用这种新方法，我们在不破坏生物样本的情况下进行了测量。我们计算了 dox 从细胞外部分转移到细胞内的速率常数以及 dox 在活细胞中代谢为 dox'ol 的速率。对于光谱显微镜和 LC/MS 方法，dox 进入单个细胞的转运速率相似（~1.5 pM min cell）。与光谱显微镜相比，使用 LC/MS 方法时，代谢速率大约每 500 个细胞就会低估一次。显微镜方法进一步表明，dox 和 dox'ol 以不同的速率（分别为 0.8 和 0.3 pM min）转运到核内。LC/MS 不是确定药物从细胞质转运到核内的实际方法。我们使用各种方法证实，当与高分辨率成像结合使用时，分子的光谱特征可以用作分析药物代谢的有力方法。我们提出，光谱显微镜是一种新的方法，可以在单细胞水平上研究药物定位、转运、转化和鉴定，而 LC/MS 更适合在器官或组织水平上进行药物筛选。
