聚乙二醇400对黄芩苷及其主代谢物胆汁排泄的影响

Effect of Polyethylene Glycol 400 on Bile Excretion of Baicalin and Its Main Metabolite

目的:考察药用辅料聚乙二醇400(PEG400)对黄芩苷及其主代谢物黄芩素6-O-β-D-葡萄糖醛酸苷(B6G)胆汁排泄的影响,并分析其作用机制。方法:大鼠随机分为黄芩苷+水组与黄芩苷+PEG400组,利用10%水合氯醛(剂量4 mL·kg-1)腹腔注射诱导麻醉,制作大鼠胆管插管模型,待大鼠完全清醒后,以168 mg·kg-1剂量的黄芩苷分别给大鼠灌胃相应的黄芩苷水溶液和黄芩苷PEG400溶液,收集给药后0~12 h的胆汁,使用UPLC-MS/MS测定不同时间段药物经胆汁排泄的浓度,采用Thermo Hypersil GOLD C18色谱柱(2.1 mm×100 mm,1.9μm),流动相乙腈(A)-0.1%甲酸水溶液(B)梯度洗脱(0~9 min,90%~27%B;9~10 min,27%~90%B;10~12 min,90%B),流速0.3 mL·min^-1,柱温30℃,进样量5μL;质谱条件为电喷雾离子源(ESI),正离子检测模式。利用体外孵育法和酶联免疫吸附测定(ELISA)研究PEG400对尿苷二磷酸葡萄糖醛酸转移酶(UGT)1A8和UGT1A9活性及二者在大鼠肝脏中表达的影响。结果:与黄芩苷+水组相比,黄芩苷+PEG400组大鼠12 h内胆汁中黄芩苷及其主代谢物B6G的累积排泄量分别增加了1.8,2.1倍;PEG400分别使UGT1A8与UGT1A9的酶活性提高了2.0,1.5倍,使肝脏中UGT1A8与UGT1A9的质量分数提高了2.2,1.3倍。结论:PEG400可通过提高UGT1A8与UGT1A9的活性和表达来显著增加黄芩苷及其主代谢物B6G的胆汁排泄,且对B6G胆汁排泄的促进作用大于原形药物黄芩苷,可为PEG400的临床合理应用及黄芩苷等黄酮类药物新剂型的设计提供依据。

Objective:To investigate the effect of polyethylene glycol 400(PEG400)on rat bile excretion of baicalin and its main metabolite[baicalein 6-O-β-D-glucuronide(B6G)],and to analyze its mechanism of action.Method:Rats were randomly divided into baicalin+water group and baicalin+PEG400 group,the anesthesia was induced by intraperitoneal injection of 10%chloral hydrate(dose of 4 mL·kg-1)to prepare a rat bile duct intubation model.After the rats were fully awake,rats were given baicalin aqueous solution and baicalin PEG400 solution with dose of 168 mg·kg-1 for baicalin,respectively.And bile was collected from 0 h to 12 h after administration.UPLC-MS/MS was used to determine the concentration of drug excreted through bile at different time periods.Thermo Hypersil GOLD C18 column was used with acetonitrile(A)-0.1%formic acid solution(B)as the mobile phase for gradient elution(0-9 min,90%-27%B;9-10 min,27%-90%B;10-12 min,90%B),the flow rate was 0.3 mL·min^-1,the column temperature was 30℃,the injection volume was 5μL.The mass spectra were obtained in positive ion mode with electrospray ionization(ESI).The effects of PEG400 on the activities and expressions in rat liver of uridine diphosphate glucuronyltransferase(UGT)1A8 and UGT1A9 were studied in vitro incubation assay and enzyme linked immunosorbent assay(ELISA).Result:Compared with the baicalin+water group,in the baicalin+PEG400 group,the bile cumulative excretions of baicalin and B6G increased by 1.8 times and 2.1 times within 12 h,respectively.PEG400 increased the enzyme activities of UGT1A8 and UGT1A9 by 2.0 times and 1.5 times,and their concentrations in liver were increased by 2.2 times and 1.3 times,respectively.Conclusion:PEG400 can significantly increase the bile excretion of baicalin and its main metabolite B6G by enhancing the activities and expressions of UGT1A8 and UGT1A9,and its promoting effect on bile excretion of B6G is greater than that of baicalin,which provides a basis for the rational clinical application of PEG400 and the design of new dosage forms of flavonoids such as baicalin.