Abstract：
BACKGROUND: Rhizoma Coptidis (RC), the dried rhizome of Coptis Chinensis Franch., can dispel dampness and heat within the body and has been traditionally used for the treatment of cardiovascular disease (CVD)-associated problems including hyperlipidemia in China. Berberine (BBR) is the main active component of RC, which has been shown to possess significant therapeutic potential. However, only 0.14% of BBR is metabolized in the liver, and the extremely low bioavailability (<1%) and blood concentration of BBR in experimental and clinical settings is insufficient to achieve the effects as observed under in vitro conditions, which imposes challenges to explain its excellent pharmacological actions. Intense efforts are currently being devoted to defining its specific pharmacological molecular targets, while the exploration from the perspective of its pharmacokinetic disposition has rarely been reported to date, which could hardly make a comprehensive understanding of its hypolipidemic enigma.
OBJECTIVE: This study made a pioneering endeavor to unveil the hypolipidemic mechanism of BBR from RC focusing on its unique intestines-erythrocytes-mediated bio-disposition.
METHODS: The fate of BBR in intestines and erythrocytes was probed by a rapid and sensitive LC/MS-IT-TOF method. To analyze the disposition of BBR, a reliable HPLC method was subsequently developed and validated for simultaneous determination of BBR and its key active metabolite oxyberberine (OBB) in whole blood, tissues, and excreta. Meanwhile, the enterohepatic circulation (BDC) of BBR and OBB was verified by bile duct catheterization rats. Finally, lipid overloading models of L02 and HepG2 cells were employed to probe the lipid-lowering activity of BBR and OBB at in vivo concentration.
RESULTS: The results showed that BBR underwent biotransformation in both intestines and erythrocytes, and converted into the major metabolite oxyberberine (OBB). The AUC0-t ratio of total BBR to OBB was approximately 2:1 after oral administration. Besides, the AUC0-t ratio of bound BBR to its unbound counterpart was 4.6:1, and this ratio of OBB was 2.5:1, indicative of abundant binding-type form in the blood. Liver dominated over other organs in tissue distribution. BBR was excreted in bile, while the excretion of OBB in feces was significantly higher than that in bile. Furthermore, the bimodal phenomenon of both BBR and OBB disappeared in BDC rats and the AUC0-t was significantly lower than that in the sham-operated control rats. Interestingly, OBB significantly decreased triglycerides and cholesterol levels in lipid overloading models of L02 and HepG2 cells at in vivo-like concentration, which was superior to the prodrug BBR.
CONCLUSIONS: Cumulatively, BBR underwent unique extrahepatic metabolism and disposition into OBB by virtue of intestines and erythrocytes. BBR and OBB were mainly presented and transported in the protein-bound form within the circulating erythrocytes, potentially resulting in hepatocyte targeting accompanied by obvious enterohepatic circulation. The unique extrahepatic disposition of BBR via intestines and erythrocytes conceivably contributed enormously to its hypolipidemic effect. OBB was the important material basis for the hypolipidemic effect of BBR and RC.
OBJECTIVE: This study made a pioneering endeavor to unveil the hypolipidemic mechanism of BBR from RC focusing on its unique intestines-erythrocytes-mediated bio-disposition.
METHODS: The fate of BBR in intestines and erythrocytes was probed by a rapid and sensitive LC/MS-IT-TOF method. To analyze the disposition of BBR, a reliable HPLC method was subsequently developed and validated for simultaneous determination of BBR and its key active metabolite oxyberberine (OBB) in whole blood, tissues, and excreta. Meanwhile, the enterohepatic circulation (BDC) of BBR and OBB was verified by bile duct catheterization rats. Finally, lipid overloading models of L02 and HepG2 cells were employed to probe the lipid-lowering activity of BBR and OBB at in vivo concentration.
RESULTS: The results showed that BBR underwent biotransformation in both intestines and erythrocytes, and converted into the major metabolite oxyberberine (OBB). The AUC0-t ratio of total BBR to OBB was approximately 2:1 after oral administration. Besides, the AUC0-t ratio of bound BBR to its unbound counterpart was 4.6:1, and this ratio of OBB was 2.5:1, indicative of abundant binding-type form in the blood. Liver dominated over other organs in tissue distribution. BBR was excreted in bile, while the excretion of OBB in feces was significantly higher than that in bile. Furthermore, the bimodal phenomenon of both BBR and OBB disappeared in BDC rats and the AUC0-t was significantly lower than that in the sham-operated control rats. Interestingly, OBB significantly decreased triglycerides and cholesterol levels in lipid overloading models of L02 and HepG2 cells at in vivo-like concentration, which was superior to the prodrug BBR.
CONCLUSIONS: Cumulatively, BBR underwent unique extrahepatic metabolism and disposition into OBB by virtue of intestines and erythrocytes. BBR and OBB were mainly presented and transported in the protein-bound form within the circulating erythrocytes, potentially resulting in hepatocyte targeting accompanied by obvious enterohepatic circulation. The unique extrahepatic disposition of BBR via intestines and erythrocytes conceivably contributed enormously to its hypolipidemic effect. OBB was the important material basis for the hypolipidemic effect of BBR and RC.
摘要：
背景：黄连（RC），黄连干根茎。,可以消除体内的湿热，传统上已用于治疗心血管疾病（CVD）相关问题，包括中国的高脂血症。小檗碱（BBR）是RC的主要活性成分，已被证明具有显著的治疗潜力。然而,只有0.14%的BBR在肝脏中代谢，和极低的生物利用度（<1％）和血液浓度的BBR在实验和临床设置是不够的，以实现在体外条件下观察到的效果，这对解释其优异的药理作用提出了挑战。目前正在为确定其特定的药理分子靶标而付出巨大的努力，虽然迄今为止很少报道从其药代动力学处置的角度进行探索，这很难对其降血脂之谜有全面的了解。
目的：本研究从RC揭示了BBR的降血脂机制，重点是其独特的肠-红细胞介导的生物配置。
方法：用快速、灵敏的LC/MS-IT-TOF方法检测肠和红细胞中BBR的命运。为了分析BBR的配置，随后开发并验证了一种可靠的HPLC方法，用于同时测定全血中BBR及其关键活性代谢产物羟小檗碱（OBB），组织，和排泄物.同时,通过胆管插管大鼠证实BBR和OBB的肝肠循环（BDC）。最后,采用L02和HepG2细胞的脂质过载模型来探测BBR和OBB在体内浓度下的降脂活性。
结果：结果显示，BBR在肠和红细胞中都进行了生物转化，并转化为主要代谢产物氧小檗碱(OBB)。口服给药后,总BBR与OBB的AUC0-t比率约为2:1。此外,结合BBR与其未结合对应物的AUC0-t比率为4.6:1,OBB的这一比率为2.5:1,表明血液中存在丰富的结合型形式.肝脏在组织分布上优于其他器官。BBR在胆汁中排泄,而粪便中OBB的排泄量明显高于胆汁。此外，BDC大鼠BBR和OBB双峰现象均消失，AUC0-t明显低于假手术对照大鼠。有趣的是,OBB在体内样浓度下显著降低L02和HepG2细胞的脂质超负荷模型中的甘油三酯和胆固醇水平,优于前药BBR。
结论：累积，BBR凭借肠和红细胞经历了独特的肝外代谢和向OBB的处置。BBR和OBB主要在循环红细胞内以蛋白结合形式存在和转运，可能导致肝细胞靶向并伴有明显的肝肠循环。可以想象，BBR通过肠和红细胞的独特肝外分布极大地促进了其降血脂作用。OBB是BBR和RC降血脂作用的重要物质基础。
目的：本研究从RC揭示了BBR的降血脂机制，重点是其独特的肠-红细胞介导的生物配置。
方法：用快速、灵敏的LC/MS-IT-TOF方法检测肠和红细胞中BBR的命运。为了分析BBR的配置，随后开发并验证了一种可靠的HPLC方法，用于同时测定全血中BBR及其关键活性代谢产物羟小檗碱（OBB），组织，和排泄物.同时,通过胆管插管大鼠证实BBR和OBB的肝肠循环（BDC）。最后,采用L02和HepG2细胞的脂质过载模型来探测BBR和OBB在体内浓度下的降脂活性。
结果：结果显示，BBR在肠和红细胞中都进行了生物转化，并转化为主要代谢产物氧小檗碱(OBB)。口服给药后,总BBR与OBB的AUC0-t比率约为2:1。此外,结合BBR与其未结合对应物的AUC0-t比率为4.6:1,OBB的这一比率为2.5:1,表明血液中存在丰富的结合型形式.肝脏在组织分布上优于其他器官。BBR在胆汁中排泄,而粪便中OBB的排泄量明显高于胆汁。此外，BDC大鼠BBR和OBB双峰现象均消失，AUC0-t明显低于假手术对照大鼠。有趣的是,OBB在体内样浓度下显著降低L02和HepG2细胞的脂质超负荷模型中的甘油三酯和胆固醇水平,优于前药BBR。
结论：累积，BBR凭借肠和红细胞经历了独特的肝外代谢和向OBB的处置。BBR和OBB主要在循环红细胞内以蛋白结合形式存在和转运，可能导致肝细胞靶向并伴有明显的肝肠循环。可以想象，BBR通过肠和红细胞的独特肝外分布极大地促进了其降血脂作用。OBB是BBR和RC降血脂作用的重要物质基础。
