金钗石斛细粉与超微粉对大鼠肠道转运体影响的比较
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摘要
目的:比较金钗石斛细粉和超微粉中生物碱类成分在大鼠体内的血药浓度,考察不同粒径金钗石斛对肠道转运体基因表达的影响。方法:将36只大鼠随机分为空白组、金钗石斛细粉组(0.25 g·kg~(-1))和金钗石斛超微粉组(0.25 g·kg~(-1)),每6 h灌胃1次,连续灌胃5 d,收集大鼠血浆和小肠样本。血浆样本分析采用UPLC-MS,Hypersil Gold C_(18)色谱柱(2.1 mm×150 mm,1.9μm),以0.1%甲酸水溶液-乙腈为流动相进行梯度洗脱;电喷雾离子源(ESI)正离子模式检测。采用实时荧光定量聚合酶链式反应分析肠道转运体多药耐药蛋白1(MDR1),寡肽转运蛋白1(PEPT1),有机阳离子转运蛋白2(OCT2),乳腺癌耐药蛋白1(BCRP1),单羧酸转运蛋白1(MCT1)和多药耐药相关蛋白2(MRP2)的基因表达情况。结果:灌胃给予金钗石斛细粉和超微粉后,在大鼠血浆中能检测出石斛碱、石斛氨碱和mubironine B。金钗石斛超微粉组血浆中石斛碱和石斛氨碱的质量浓度比金钗石斛细粉组显著增加(P<0.05)。与空白组比较,金钗石斛细粉和超微粉均能显著抑制BCRP1的基因表达(P<0.05),且金钗石斛超微粉组较细粉组表达更低(P<0.05);金钗石斛细粉还能显著上调MDR1的基因表达(P<0.05),而金钗石斛超微粉则对MDR1基因表达的影响不显著。结论:与金钗石斛细粉相比,金钗石斛超微粉灌胃后在大鼠体内石斛碱和石斛氨碱的血药浓度升高,这可能与肠道转运体MDR1和BCRP1有关,为金钗石斛药材粒径的选择提供了实验依据。
Objective:To compare the contents of alkaloids from fine and ultrafine powder of Dendrobium nobile stem in rat plasma,and investigate the effect of D.nobile stem with different particle sizes on gene expression of intestinal transporters.Method:Rats were randomly divided into the blank group,fine powder group of D.nobile stem(0.25 g·kg~(-1))and ultrafine powder group of D.nobile stem(0.25 g·kg~(-1)).The rats were gavaged every 6 h for 5 days.The samples of rat plasma and small intestine were collected.The plasma samples were detected with UPLC-MS.The chromatography separation was performed on a Hypersil Gold C_(18)column(2.1 mm×150 mm,1.9μm)with acetonitrile-0.1%formic acid solution as mobile phase for gradient elution.Electrospray ionization(ESI)was applied and operated in positive ion mode.The mRNA expression of multidrug resistance protein 1(MDR1),oligopeptide transporter protein 1(PEPT1),organic cation transporter protein 2(OCT2),breast cancer resistance protein 1(BCRP1),monocarboxylate transport protein 1(MCT1)and multidrug resistance related protein 2(MRP2)in small intestine were quantified by real time fluorescence quantitative polymerase chain reaction.Result:After intragastric administration of fine and ultrafine powder of D.nobile stem,dendrobine,mubironine B and dendramine could be detected in rat plasma.The contents of dendrobine and dendramine in the ultrafine powder group were significantly higher than that in the fine powder group(P<0.05).Compared with the blank group,gene expression of BCRP1 was significantly inhibited by fine and ultrafine powder of D.nobile stem(P<0.05),and the expression in the ultrafine powder group was lower than that in the fine powder group(P<0.05).The gene expression of MDR1 was up-regulated by fine powder of D.nobile stem(P<0.05).Conclusion:Compared with the fine powder group of D.nobile stem,the plasma concentrations of dendrobine and dendramine in the ultrafine powder group are significantly increased,it may be related to the intestinal transporters of MDR1 and BCRP1.These results can provide experimental basis for selecting particle size of D.nobile stem.
Objective:To compare the contents of alkaloids from fine and ultrafine powder of Dendrobium nobile stem in rat plasma,and investigate the effect of D.nobile stem with different particle sizes on gene expression of intestinal transporters.Method:Rats were randomly divided into the blank group,fine powder group of D.nobile stem(0.25 g·kg~(-1))and ultrafine powder group of D.nobile stem(0.25 g·kg~(-1)).The rats were gavaged every 6 h for 5 days.The samples of rat plasma and small intestine were collected.The plasma samples were detected with UPLC-MS.The chromatography separation was performed on a Hypersil Gold C_(18)column(2.1 mm×150 mm,1.9μm)with acetonitrile-0.1%formic acid solution as mobile phase for gradient elution.Electrospray ionization(ESI)was applied and operated in positive ion mode.The mRNA expression of multidrug resistance protein 1(MDR1),oligopeptide transporter protein 1(PEPT1),organic cation transporter protein 2(OCT2),breast cancer resistance protein 1(BCRP1),monocarboxylate transport protein 1(MCT1)and multidrug resistance related protein 2(MRP2)in small intestine were quantified by real time fluorescence quantitative polymerase chain reaction.Result:After intragastric administration of fine and ultrafine powder of D.nobile stem,dendrobine,mubironine B and dendramine could be detected in rat plasma.The contents of dendrobine and dendramine in the ultrafine powder group were significantly higher than that in the fine powder group(P<0.05).Compared with the blank group,gene expression of BCRP1 was significantly inhibited by fine and ultrafine powder of D.nobile stem(P<0.05),and the expression in the ultrafine powder group was lower than that in the fine powder group(P<0.05).The gene expression of MDR1 was up-regulated by fine powder of D.nobile stem(P<0.05).Conclusion:Compared with the fine powder group of D.nobile stem,the plasma concentrations of dendrobine and dendramine in the ultrafine powder group are significantly increased,it may be related to the intestinal transporters of MDR1 and BCRP1.These results can provide experimental basis for selecting particle size of D.nobile stem.
引文
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[5]黄琦,廖鑫,吴芹,等.金钗石斛生物总碱对糖尿病大鼠血糖及肝脏组织IRS-2 mRNA,IGF-1 mRNA表达的影响[J].中国实验方剂学杂志,2014,20(19):155-158.
[6]陈志国,叶松山,范迎,等.金钗石斛多糖提取工艺的优化及对小鼠脾细胞增殖的影响[J].中国实验方剂学杂志,2011,17(15):27-32.
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[8]梁勇.纳米微粒在医药学中的应用[J].中国粉体技术,2001,7(5):39-41.
[9]贺雨馨,曾宇馨,祝天添,等.金钗石斛超微粉增强免疫力作用研究[J].现代中药研究与实践,2018,32(3):18-21.
[10]Ng T B,LIU J,Wong J H,et al. Review of research on Dendrobium,a prized folk medicine[J]. Appl Microbiol Biotechnol,2012,93(5):1795-1803.
[11]何芋岐,鲁艳柳,李利生,等.基于UPLC-ESI-OrbitrapMS技术对金钗石斛生物碱的分析[J].中国实验方剂学杂志,2017,23(20):30-35.
[12]鲁艳柳,黄思,汪敏,等. UPLC/LTQ-Orbitrap-MS技术鉴定石斛碱在人肝微粒体中的代谢产物[J].中国实验方剂学杂志,2018,24(4):72-77.
[13]鲁艳柳,刘浩,曾瑶,等.石斛碱在小鼠体内的组织分布[J].中国实验方剂学杂志,2018,24(18):134-139.
[14]伊秀林,司端运,刘昌孝.应用药物转运体的药代动力学评价[J].药物评价研究,2010,33(5):341-346.
[15]Dobson P D,Kell D B. Carrier-mediated cellular uptake of pharmaceutical drugs:an exception or the rule[J].Nat Rev Drug Discov,2008,7(3):205-220.
[16]Dobson P D,Lanthaler K,Oliver S G,et al. Implications of the dominant role of transporters in drug uptake by cells[J]. Curr Top Med Chem,2009,9(2):163.
[17]刘志浩,刘克辛.肠道药物转运体及其研究方法[J].药学学报,2011,46(4):370-376.
[18]匡健,黄鑫,江振洲,等.肠道药物转运体对药物吸收的研究进展[J].药学与临床研究,2015,23(3):279-282.
[2]张晓敏,孙志蓉,陈龙,等.金钗石斛的化学成分和药理作用研究进展[J].中国现代应用药学,2014,31(7):895-899.
[3]YE Q, QIN G, ZHAO W. Immunomodulatory sesquiterpene glycosides from Dendrobium nobile[J].Phytochemistry,2002,61(8):885-890.
[4]柴金珍,黄娟萍,刘静,等.不同石斛的药理作用研究现状[J].中成药,2013,35(12):2725-2730.
[5]黄琦,廖鑫,吴芹,等.金钗石斛生物总碱对糖尿病大鼠血糖及肝脏组织IRS-2 mRNA,IGF-1 mRNA表达的影响[J].中国实验方剂学杂志,2014,20(19):155-158.
[6]陈志国,叶松山,范迎,等.金钗石斛多糖提取工艺的优化及对小鼠脾细胞增殖的影响[J].中国实验方剂学杂志,2011,17(15):27-32.
[7]舒朝晖,刘根凡,马孟骅,等.中药超微粉碎之浅析[J].中国中药杂志,2004,29(9):823-827.
[8]梁勇.纳米微粒在医药学中的应用[J].中国粉体技术,2001,7(5):39-41.
[9]贺雨馨,曾宇馨,祝天添,等.金钗石斛超微粉增强免疫力作用研究[J].现代中药研究与实践,2018,32(3):18-21.
[10]Ng T B,LIU J,Wong J H,et al. Review of research on Dendrobium,a prized folk medicine[J]. Appl Microbiol Biotechnol,2012,93(5):1795-1803.
[11]何芋岐,鲁艳柳,李利生,等.基于UPLC-ESI-OrbitrapMS技术对金钗石斛生物碱的分析[J].中国实验方剂学杂志,2017,23(20):30-35.
[12]鲁艳柳,黄思,汪敏,等. UPLC/LTQ-Orbitrap-MS技术鉴定石斛碱在人肝微粒体中的代谢产物[J].中国实验方剂学杂志,2018,24(4):72-77.
[13]鲁艳柳,刘浩,曾瑶,等.石斛碱在小鼠体内的组织分布[J].中国实验方剂学杂志,2018,24(18):134-139.
[14]伊秀林,司端运,刘昌孝.应用药物转运体的药代动力学评价[J].药物评价研究,2010,33(5):341-346.
[15]Dobson P D,Kell D B. Carrier-mediated cellular uptake of pharmaceutical drugs:an exception or the rule[J].Nat Rev Drug Discov,2008,7(3):205-220.
[16]Dobson P D,Lanthaler K,Oliver S G,et al. Implications of the dominant role of transporters in drug uptake by cells[J]. Curr Top Med Chem,2009,9(2):163.
[17]刘志浩,刘克辛.肠道药物转运体及其研究方法[J].药学学报,2011,46(4):370-376.
[18]匡健,黄鑫,江振洲,等.肠道药物转运体对药物吸收的研究进展[J].药学与临床研究,2015,23(3):279-282.
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