王不留行环肽体内代谢及降血糖构效关系研究

The worldwide research on cyclicpeptides is still in its early stage, and this kind of compounds was reported to exhibit various biological activities, including immune-suppression, antitumor, sleeping peacefully, antibacterial activity and etc. It has been found by our laboratory that cyclicpeptides from Vaccaria segetalis has significant hypoglycemic efficacy in GK rats, a model of spontaneous type 2 diabetes mellitus. SCP was purified further and then several cyclicpeptides were isolated. The study on metabolism of SCP in rats and in vitro with rat liver hepatocytes showed that SCP can be mainly metabolized by liver and the structure of metabolites is characterized by intra-molecular ring not opening, metabolic reactions occurring in the side chain, and chiral carbon number, position and stereo configuration changed. The previous study on pharmacological mechanism showed that the original compounds from SCP and their metabolites can inhibit glycogen gluconeogenesis significantly. Besides, metabolites can also promote myotubes glucose consumption activity. Our preliminary results obtained in SCP hypoglycemic effect and pharmacological mechanisms is a new discovery about biological activity of the whole cyclic peptides. At present, the hypoglycemic drugs on the market also do not have compound of this kind structure. In our subject, analysis on the metabolites of SCP, preparation of metabolites, structure modification and pharmacological mechanism will be studied systematically to clarify the structure-activity relationship，which could also provide a scientific basis to develop a new generation drugs of diabetes with cyclic peptide structure.

环肽类成分研究的历史不长，迄今报道的环肽药理活性主要是免疫抑制、抗肿瘤、安眠和抗菌等。本实验室发现王不留行环肽部位（SCP）对自发性糖尿病GK大鼠具有显著降糖药效，进一步分离得到SCP-D等多个环肽单体。我们初步研究表明，SCP主要经肝脏代谢，其代谢产物结构特点是环肽母核不开环，代谢反应发生在侧链，部分结构中的手性碳数量、位置和立体构型发生改变。初步药理机制研究表明，SCP单体原形和代谢产物均具有明显的抑制肝糖异生作用，代谢物还有促肌管细胞葡萄糖消耗的活性。我们获得的SCP降血糖作用和药理机制的初步结果，是整个环肽类结构生物活性的新发现，目前已上市的降糖药物也无此类结构。本申请将对SCP原形及代谢产物进行分析鉴定、人工合成、结构修饰和药理机制等系统研究，阐明结构与降血糖的药效关系，为研究以环肽为结构模板的新一代治疗糖尿病药物提供科学依据。

利用质谱从王不留行提取物中共鉴定出52个化合物，包括3个酚酸类成分，4个黄酮苷类成分，6个环肽类成分，37个皂苷类化合物。利用大孔吸附树脂分离得到王不留行黄酮苷部位、环肽部位和皂苷部位，发现环肽部位可降低GK大鼠空腹血糖和口服糖耐量。经过进一步植物化学分离、提取和纯化，获得4个王不留行环肽（SCP）单体，并且通过质谱、氢谱、碳谱等数据鉴定为其结构分别为SCP-A, SCP-B,SCP-D和SCP-E.同时，，还人工全合成了SCP-A, SCP-B,SCP-D,SCP-E和SCP-I 五个环肽单体。.将上述单体进行体外（原代肝细胞）和体内（大鼠含药尿）代谢实验，并利用HPLC-Q-TOF分析鉴定代谢物，最终发现5个SCP-A代谢物，5个SCP-B 代谢物、7个SCP-D代谢物和5个SCP-E 代谢物，其中芳环上的氧化代谢途径为主要代谢途径。经过药理研究，发现 SCP-A, SCP-B,SCP-D和SCP-E均可以抑制肝糖异生，这可能是环肽部位发挥降糖作用的机制之一。.此外，我们采用人工全合成的方法合成制备出具有多个手性中心的Janadolide 和Alotamide A及其多个衍生物。其中以18步反应、7.6%的总收率化合物Janadolide，并次合成了C7位构型翻转的衍生物7-epi Janadolide，总收率为7.5%。首次合成了其中4个C19位为S构型的Alotamide A非对映异构体，为(19S,28R,30S)-1-117e，(19S,28R,30R)-1-117f，(19S,28S,30R)-1-117g及(19S,28S,30S)-1-117h。通过药理筛选，发现7-epi Janadolide 和(19S,28S,30R)-1-117g可显著抑制神经元钾离子通道。