Department of Experimental Medicine, University of Campania "Luigi Vanvitelli", 80138 Naples, Italy.
Department of Mathematics and Physics, University of Campania "Luigi Vanvitelli", 81100 Caserta, Italy.
Int J Mol Sci. 2023 Aug 2;24(15):12364. doi: 10.3390/ijms241512364.
An analytical method based on tandem mass spectrometry-shotgun is presently proposed to obtain sphingolipidomic profiles useful for the characterization of lipid extract from X-ray-exposed and unexposed hepatocellular carcinoma cells (HepG2). To obtain a targeted lipidic profile from a specific biological system, the best extraction method must be identified before instrumental analysis. Accordingly, four different classic lipid extraction protocols were compared in terms of efficiency, specificity, and reproducibility. The performance of each procedure was evaluated using the Fourier-transform infrared spectroscopic technique; subsequently, the quality of extracts was estimated using electrospray ionization tandem mass spectrometry. The selected procedure based on chloroform/methanol/water was successfully used in mass spectrometry-based shotgun sphingolipidomics, allowing for evaluation of the response of cells to X-ray irradiation, the most common anticancer therapy. Using a relative quantitative approach, the changes in the sphingolipid profiles of irradiated cell extracts were demonstrated, confirming that lipidomic technologies are also useful tools for studying the key sphingolipid role in regulating cancer growth during radiotherapy.
目前提出了一种基于串联质谱- shotgun 的分析方法，用于获得有用的脂类组学图谱，以表征 X 射线照射和未照射的肝癌细胞（HepG2）的脂类提取物。为了从特定的生物系统中获得靶向脂质图谱，在进行仪器分析之前，必须确定最佳的提取方法。因此，从效率、特异性和重现性方面比较了四种不同的经典脂质提取方案。使用傅里叶变换红外光谱技术评估每个程序的性能；随后，使用电喷雾电离串联质谱法评估提取物的质量。基于氯仿/甲醇/水的选定方案成功地用于基于质谱的 shotgun 鞘脂组学，允许评估细胞对 X 射线照射的反应，这是最常见的抗癌治疗方法。使用相对定量方法，证明了照射细胞提取物中鞘脂谱的变化，证实了脂质组学技术也是研究放疗过程中关键鞘脂在调节癌症生长中的作用的有用工具。
