The Metabolism And Pharmacokinetics Of Isocorydine And Protopine In Dactylicapnos Scandens

Dactylicapnos scandens (D. Don) Hutch., a perennial herb, is mainly distributed in Yunnan, southwestern Sichuan, western Guangxi province and southeastern Tibet Autonomous Region of China. Dactylicapnos scandens, prepared from the dried tubers of Dactylicapnos scandens (D. Don) hutch, is a well-known Traditional Chinese medicine (TCM) with the Chinese name "Zijinlong". Dactylicapnos scandens is bitter and pungent in taste, cool-natured and toxic, used for the treatment of hypertension, inflammation, blooding and pain for centuries. Dactylicapnos scandens was a Bai Nationality folk medicine with significant analgesic effect, and was recorded in the Chinese Pharmacopoeia (1977version). The main chemical constituents of Dactylicapnos scandens were alkaloids, in which isocorydine (ICD) and protopine (PTP) was the most abundant, account for5.2%and1.4%of the crude drug, respectively. It was found that both isocorydine and protopine showed various pharmacological activities, such as spasmolytic, vasodilating, antiplasmodial and antiarrhythmic effects for isocorydine, and spasmolytic, antithrombotic, antiinflammatory, anticholinergic and antihistamine effects for protopine. However, the metabolism, pharmacokinetics and transport properties of isocorydine, as well as the transport of protopine, still have not been evaluated. In addition, the absorption, distribution, metabolism and excretion (ADME) of Dactylicapnos scandens was still unclear. Therefore, in order to provide the scientific basis and clinical reference for the development and utilization of Dactylicapnos scandens, the present study will investigate the in vitro and in vivo metabolism, pharmacokinetics and transport of isocorydine and protopine.1. Metabolism of isocorydine in rat liver microsomesIsocorydine could be metabolized by CYPs (Phase Ⅰ metabolism), but no Phase Ⅱ metabolites were detected in both Phase Ⅱ glucuronidation metabolism and the Sequential metabolism of phase Ⅰ and phase Ⅱ in rat liver microsomes (RLMs). CYP3A2was involved in isocorydine oxidative metabolism. Classical CYPs’ substrates were used to investigate the inhibition effect of isocorydine on the CYPs, with the results that isocorydine almost has no inhibition effect on these main CYP isoforms. Three metabolites of isocorydine in RLMs were analyzed by high resolution mass spectrometry (HRMS) as two mono-demethylation metabolites and a mono-hydroxylation metabolite.2. Metabolism of isocorydine in human liver microsomes, recombinant human CYPs and HepG2cellsIt was found that CYP2C8,2C9,2C19and CYP3A4were main CYP isoforms that mediated the metabolism of isocorydine in human. Isocorydine did not inhibit CYP1A2,2C9,2C19,2D6,2E1and3A4. Isocorydine at10μmol/L showed almost no gene induction in CYP2B6, CYP3A4and UGT1A1, assayed by dual-luciferase reporter assay. Isocorydine and protopine showed almost no toxicity on HepG2cells, indicating that both alkaloids may have insignificant hepatotoxicity.3. Transport and uptake studies of isocorydine and protopine in MDR1, BCRP and OCT1transgenic cellsTransgenic cells MDCK-MDR1, LLC-PK1-BCRP and MDCK-OCT1were adopted as the in vitro model to predict the intestinal absorption and blood-brain barrier (BBB) permeability. Isocorydine and protopine were not the substrates of P-gp, BCRP and OCT1. Isocorydine and protopine showed no inhibition on P-gp. Been not the substrates of efflux transporters P-gp and BCRP, isocorydine and protopine may have good oral bioavailability and BBB permeability. The IC50of isocorydine and protopine on PC12cells were85.5±1.0μmol/L and75.3±0.9μmol/L, respectively, demonstrating that the two alkaloids may have potential neurotoxicity.4. Pharmacokinetics and tissue distribution of isocorydine in ratsA method of analysis of isocorydine in rat plasma and tissues using LC-MS/MS was developed and validated with respect to specificity, accuracy, precision, recovery, and successfully applied to evaluate the pharmacokinetics and tissue distribution of isocorydine in rats. In the present study, the Tmax of isocorydine in plasma was<20min showed that isocorydine was rapidly absorbed, while the t1/2of isocorydine in plasma was<1h showed that isocorydine was quickly eliminated. The absolute bioavailability (F) was estimated as33.4%, indicating a good oral absorption of isocorydine. Tissue distribution studies showed that isocorydine could be rapidly and widely distributed into tissues and it can efficiently cross the blood-brain barrier in rats.5. Pharmacokinetics, tissue distribution and excretion of the extract of Dactylicapnos scandens in ratsA LC-ESI-MS/MS method for the simultaneous analysis of isocorydine and protopine in Dactylicapnos scandens in rat biological matrices was developed and applied to investigate the pharmacokinetics, tissue distribution and excretion of the Dactylicapnos scandens extract in rats. The Tmax and the t1/2of isocorydine, which is0.333±0.144h and1.965±0.387h respectively, shows that isocorydine can be rapidly absorbed and quickly eliminated in plasma. The absolute bioavailability (F) was estimated as29.1%, indicating the considerable oral absorption of isocorydine. The Tmax, t1/2a and bioavailability of protopine was2.667±1.155h,2.609±0.739h and25.8%, respectively. Tissue distribution studies showed that isocorydine and protopine could rapidly and widely distribut into tissues and can effectively cross the BBB in rats, while the excretion study indicated that very low proportion of isocorydine (<5.0%) and protopine (<1.0%) was excreted as the unchanged form. To the best of our knowledge, this is the first report for the thorough studies of the pharmacokinetics, tissue distribution and excretion profiles of Dactylicapnos scandens in rats. The acquired data would provide helpful information for the clinical applications or further development of Dactylicapnos scandens.