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目的采用马丁达比犬肾上皮(MDCK)细胞模型研究广藿香醇跨膜转运机制。方法采用Agilent气相色谱质谱联用仪测定广藿香醇浓度,色谱柱为Agilent HP-5ms(5%苯基甲基硅氧烷,30 m×0.25 mm,0.25μm),载气为氦气;质谱采用电子碰撞离子源,电子倍增电压模式,检测离子为m/z 222.2(广藿香醇)和m/z 190.1(内标异土木香内酯)。建立MDCK细胞模型,通过改变广藿香醇浓度、转运方向考察广藿香醇的跨膜转运机制;分别加入外排蛋白P-糖蛋白(P-gp)、多药耐药相关蛋白(MRP)、乳腺癌耐药蛋白(BCRP)抑制剂,考察外排转运体对广藿香醇跨膜的影响。结果广藿香醇浓度40、20和10μg·m L~(-1)的双向转运表观渗透系数(P_(app))值均大于10×10~(-5) cm·s~(-1),各剂量组不同转运方向的P_(app)比值均小于1.5。抑制转运试验中,加入P-gp抑制剂后,双向P_(app)值变化显著;加入BCRP抑制剂后,双向P_(app)值变化不显著;加入MRP抑制剂后,广藿香醇肠内壁侧到肠腔侧的P_(app)值显著降低。结论广藿香醇具有高透膜性,其跨膜转运机制以被动转运为主,同时可能有转运载体参与。P-gp和MRP可能参与到广藿香醇跨膜过程,BCRP不参与广藿香醇的转运过程。
Objective To study the mechanism of patchouli alcohol transmembrane transport by using the model of canine derby kidney (MDCK) cells. Methods The patchouli alcohol concentration was determined by Agilent Gas Chromatography-Mass Spectrometer. The column was Agilent HP-5ms (5% phenylmethylsiloxane, 30 m × 0.25 mm, 0.25 μm) with helium carrier gas. Mass spectrometry Electron impact ionization source, electron multiplying voltage mode, m / z 222.2 (patchoulol) and m / z 190.1 (internal isocorodolactone) were used as detection ions. The MDCK cell model was established. The transmembrane transport mechanism of patchouli alcohol was investigated by changing the patchouli alcohol concentration and transport direction. P-glycoprotein (P-gp), multidrug resistance-related protein (MRP) Cancer drug resistance protein (BCRP) inhibitors to investigate the effect of efflux transporters on patchouli alcohol transmembrane. Results The apparent permeability coefficients (P_ (app)) of patchouli alcohol bidirectional transport at the concentrations of 40, 20 and 10 μg · m L -1 were higher than 10 × 10 -5 cm · s -1, The P_ (app) ratios in different transport directions of each dose group were less than 1.5. Inhibition of transport test, after adding P-gp inhibitor, two-way P_ (app) value changes significantly; BCRP inhibitor added, two-way P_ (app) values did not change significantly; adding MRP inhibitor, patchouli alcohol intestinal wall side P app values to the luminal side were significantly lower. Conclusions Patchouli alcohol has high permeability and its transmembrane transport mechanism is mainly passive transport, meanwhile transporter may be involved. P-gp and MRP may participate in the patchouli alcohol transmembrane process, BCRP is not involved in patchouli alcohol transport process.