Xiao Si-Jia, Li Shan-Shan, Xie Bin, Chen Wei, Xu Xi-Ke, Zu Xian-Peng, Shen Yun-Heng
Department of Natural Medicinal Chemistry, School of Pharmacy, Naval Medical University Shanghai 200433 China
School of Pharmaceutical Sciences, Yunnan Key Laboratory of Pharmacology for Natural Products, Kunming Medical University Kunming 650500 Yunnan China.
RSC Adv. 2021 Nov 23;11(60):37752-37759. doi: 10.1039/d1ra07915h.
Ingenol, as the precursor of the marketed drug ingenol mebutate, has been proven to have a variety of bioactivities. The purpose of this study was to identify the metabolites of ingenol using ultra-performance liquid chromatography-quadrupole time-of-flight-mass spectrometry (UPLC-Q/TOF-MS) combined with UNIFI software. Plasma, urine and fecal samples of rats were obtained and analyzed. A total of 18 metabolites were detected and identified in rat, including five phase II metabolites (M14-M18). Moreover, as microbial biotransformation is helpful to obtain sufficient reference standards of metabolites, the co-culture of ingenol with the fungus bio-110930 was also studied and yielded 4 phase I metabolites, in which reference standards of three metabolites were further obtained by preparative scale biotransformation. By matching their retention times, accurate masses, and fragment ions with metabolites in rat, the structures of three metabolites (M2, M3 and M4) were unambiguously confirmed by NMR technology. The results revealed that bio-110930 functioned as an appropriate model to mimic and prepare phase I metabolism of ingenol to a certain extent. It also revealed that hydroxylation, oxygenation, sulfonation, and glucuronidation were the major metabolic pathways of ingenol. Furthermore, the first systematic metabolic study of ingenol is of great significance to elucidate the metabolites and metabolic pathways , which is helpful to predict metabolites of ingenol in humans, understand the elimination mechanism of ingenol, and clarify its effectiveness and toxicity.
作为市售药物 ingenol mebutate 的前体， ingenol 已被证明具有多种生物活性。本研究的目的是使用超高效液相色谱 - 四极杆飞行时间质谱联用仪（UPLC-Q/TOF-MS）结合 UNIFI 软件来鉴定 ingenol 的代谢产物。获取并分析了大鼠的血浆、尿液和粪便样本。在大鼠中总共检测并鉴定出 18 种代谢产物，包括 5 种 II 相代谢产物（M14 - M18）。此外，由于微生物生物转化有助于获得足够的代谢产物参考标准品，还研究了 ingenol 与真菌 bio-110930 的共培养，并产生了 4 种 I 相代谢产物，其中通过制备规模生物转化进一步获得了 3 种代谢产物的参考标准品。通过将它们的保留时间、精确质量和碎片离子与大鼠中的代谢产物进行匹配，利用核磁共振技术明确确认了 3 种代谢产物（M2、M3 和 M4）的结构。结果表明，bio-110930 在一定程度上可作为模拟和制备 ingenol I 相代谢的合适模型。还表明羟基化、氧化、磺化和葡萄糖醛酸化是 ingenol 的主要代谢途径。此外，ingenol 的首次系统代谢研究对于阐明代谢产物和代谢途径具有重要意义，这有助于预测 ingenol 在人体内的代谢产物，了解 ingenol 的消除机制，并阐明其有效性和毒性。
