甲基莲心碱对大鼠肝CYP450酶含量及CYP2D1, CYP3A1,CYP2E1 mRNA的影响

目的 :探讨甲基莲心碱对大鼠微粒体混悬液蛋白中细胞色素P450(CYP450)含量,及其对CYP450亚型CYP3A1,CYP2D1,CYP2E1的mRNA表达水平的影响。 方法 :Wistar大鼠48只,随机分成空白组、肝药酶诱导剂组、肝药酶抑制剂组,甲基莲心碱(Nef)低、中、高剂量组。对照组给予1%羧甲基纤维素钠(CMC-Na),诱导剂组地塞米松100 mg ·kg^-1,抑制剂组酮康唑40 mg ·kg^-1,Nef低、中、高剂量组分别为Nef 10,20, 50 mg ·kg^-1 灌胃给药,1日1次,连续6 d。取大鼠的肝制备肝微粒体,测定肝微粒体中的蛋白浓度,用分光光度计对样品进行比色测定CYP450总酶的含量。并采用实时定量反转录——聚合酶链反应(Quantitive real-time RT-PCR)定量分析了大鼠CYP450亚型CYP3A1,CYP2D1,CYP2E1的mRNA表达水平。 结果 :甲基莲心碱在高剂量组的P450含量与空白组比较有显著性差异(P＜0.01),提示该药在较大剂量时(20~50 mg ·kg^-1)对CYP450有诱导作用。Nef在高剂量时,分别对CYP3A1和CYP2D1有诱导作用,可显著升高二者的活性(分别为空白组的1.92,2.99倍,P＜0.05);中剂量时对CYP2D1具有诱导作用(为空白组的2.40倍,P＜0.05);Nef在3个剂量下对CYP2E1均无诱导作用。 结论 :Nef在20,50 mg ·kg^-1剂量可增加CYP450总酶活性和CYP2D6 mRNA的表达,在50 mg ·kg^-1剂量可增加CYP3A1 mRNA的表达,提示Nef在较高剂量时可诱导肝药酶加速自身代谢。     

莪术油对大鼠肝微粒体CYP1A2酶活性的影响

目的:建立非那西丁及其代谢产物的液质联用检测法,研究莪术油对大鼠肝微粒体CYP1A2酶活性的影响,为临床合理用药提供参考。方法:色谱柱为XDB-C18(150mm×2.1mm,5μm),流动相为乙腈-0.1%甲酸,流速:0.4mL.min-1,柱温:30℃,以多反应监测方式采集数据。以非那西丁为探针药物,采用体外实验,实验组给予莪术油,对照组给予生理盐水,评价药物代谢酶CYP1A2酶活性的变化。结果:非那西丁和对乙酰氨基酚的检测浓度线性范围分别为4～1600ng.mL-1(r=0.9973)、3-2000ng.mL-1(r=0.9973)。实验组测得的扑热息痛/非那西丁的比值:11.30±0.71,对照组:9.60±1.04,t检验显示P<0.05,有统计学意义。结论:本法可用于检测大鼠肝微粒体孵育液中非那西丁及其代谢物浓度。莪术油对大鼠肝CYP1A2酶的活性,有体外诱导作用。     

壮骨关节丸对大鼠肝微粒体P450的影响

[目的]考察壮骨关节丸对大鼠肝微粒体细胞色素P450的影响。[方法]壮骨关节丸及其两个新工艺按含生药2.1 g/kg连续给大鼠灌胃7 d,末次药后24 h断头处死大鼠并取肝脏,用钙沉降法制备肝微粒体。在体外用含氨苯砜、非那西丁、奥美拉唑、氯唑沙宗的cocktail探针代谢来考察肝微粒体CYP3A、CYP1A2、CYP2C19、CYP2E1的活性。[结果]cocktail探针在体外肝微粒体系统中代谢1 h后,包括壮骨关节丸及其两个新工艺的给药组剩余氯唑沙宗浓度显著高于对照组,而奥美拉唑、非那西丁、氨苯砜浓度与对照组之间差异无统计学意义。[结论]壮骨关节丸可以抑制大鼠肝脏CYP2E1活性,但对CYP1A2、CYP3A及CYP2C19无显著影响。     

Inhibitory and inductive effects of Corydalis saxicola Bunting total alkaloids (CSBTA) on cytochrome P450s in rats

Corydalis saxicola Bunting, a well‐known traditional Chinese medicine in south China, has been widely used for the treatment of various hepatic diseases. Its active ingredients are Corydalis saxicola Bunting total alkaloids (CSBTA), which primarily include dehydrocavidine, palmatine, and berberine. These representative alkaloids could be metabolized by hepatic CYP450s. Hence, it is necessary to investigate the potential influences of CSBTA on CYP450s to explore the possibility of herb–drug interactions. In present study, in vitro inhibition and in vivo induction studies were performed to evaluate the potential effects of CSBTA extract on CYP450s in rats. Inhibition assay illustrated that CSBTA exerted inhibitory effects on CYP1A2 (IC₅₀, 38.08 μg/ml; K_i, 14.3 μg/ml), CYP2D1 (IC₅₀, 20.89 μg/ml; K_i, 9.34 μg/ml), CYP2C6/11 (IC₅₀ for diclofenac and S‐mephenytoin, 56.98 and 31.59 μg/ml; K_i, 39.0 and 23.8 μg/ml), and CYP2B1 (IC₅₀, 48.49 μg/ml; K_i, 36.3 μg/ml) in a noncompetitive manner. Induction study showed CSBTA had obvious inhibitory rather than inductive effects on CYP1A2 and CYP2C6/11. Interestingly, neither inhibition nor induction on CYP3A was observed for CSBTA. In conclusion, CSBTA–drug interactions might occur through CYP450s inhibition, particularly CYP1A and CYP2D. Further studies are still needed to elucidate the underlying mechanisms of inhibition.     

Inhibition of human P450 enzymes by natural extracts used in traditional medicine

Different medicinal plants are widely used in Cuba and Mexico to treat several disorders. This paper reports in vitro inhibitory effects on the P450 system of herbal products commonly used by people in Cuba and Mexico in traditional medicine for decades. Experiments were conducted in human liver microsomes. The catalytic activities of CYP1A1/2, 2D6, and 3A4 were measured using specific probe substrates. The Heliopsis longipes extract exhibited a concentration-dependent inhibition of the three enzymes, and similar effects were produced by affinin (an alkamide isolated from the H. longipes extract) and two catalytically reduced alkamides. Mangifera indica L. and Thalassia testudinum extracts, two natural polyphenol-rich extracts, diminished CYP1A1/2 and 3A4 activities, but not the CYP2D6 activity. These results suggest that these herbs inhibit the major human P450 enzymes involved in drug metabolism and could induce potential herbal-drug interactions.     

Cocktail探针药物法评价百解胶囊对肝药酶亚型CYP2C19、CYP2E1的影响

目的:研究百解胶囊对大鼠肝药酶CYP2C19、CYP2E1活性的影响,探讨其解药物毒作用的机制。方法:将大鼠随机分为4组,即空白对照组、百解胶囊组(2.43g生药/kg)、百解胶囊组(0.27g生药/kg)、苯巴比妥钠组(10.8mg/kg),按上述剂量灌胃给药。以甲硝唑为内标,建立HPLC方法检测Cocktail探针药物奥美拉唑和氯唑沙宗在大鼠体内的代谢情况,评价百解胶囊对肝药酶CYP450的影响。结果:与空白对照组比较,百解胶囊组(2.43g生药/kg)对奥美拉唑的清除率(CL/F)明显增强,曲线下面积(AUC)明显减少,其半衰期(t1/2)亦有减少趋势;百解胶囊组(2.43g生药/kg)和百解胶囊组(0.27g生药/kg)对氯唑沙宗的清除率(CL/F)明显增强,曲线下面积(AUC)明显减少,半衰期(t1/2)明显缩短。结论:百解胶囊对大鼠肝药酶CYP2C19、CYP2E1具有诱导作用,可能是其解药物毒作用的机制之一。     

参麦注射液对大鼠细胞色素P450酶亚型活性的影响

目的用Cocktail探针药物法研究参麦注射液对大鼠细胞色素P450酶(CYP450)6种亚型活性的影响。方法将SD大鼠随机分组,实验组ip给予参麦注射液(10 m L/kg),对照组ip给予等量生理盐水,诱导7 d,分别以非那西丁、安非他酮、甲苯磺丁脲、奥美拉唑、美托洛尔和咪达唑仑作为CYP1A2、CYP2B1、CYP2C9、CYP2C19、CYP2D6和CYP3A4的探针药物。UPLC-MS/MS法检测大鼠血浆中探针药物的血药浓度,采用DAS3.0软件估算药动学参数。结果与对照组相比,非那西丁、安非他酮和奥美拉唑的AUC0~∞、CL和Cmax显著降低(P0.05),甲苯磺丁脲、美托洛尔和咪达唑仑的AUC0~∞、CL和Cmax无显著性差异。结论参麦注射液对大鼠CYP1A2、CYP2B1和CYP2C19亚型的活性有明显的抑制作用,而对CYP2C9、CYP2D6和CYP3A4亚型的活性无显著性影响。     

Effects of Panax notoginseng saponins on the activities of CYP1A2, CYP2C9, CYP2D6 and CYP3A4 in rats in vivo

The aim of this study was to assess the influence of the Panax notoginseng saponins (PNS) on the activities of the drug‐metabolizing enzymes cytochrome P450 (CYP450) 1A2, 2 C9, 2D6 and 3A4 in rats. The activities of CYP1A2, 2 C9, 2D6 and 3A4 were measured using specific probe drugs. After pretreatment for 1 week with PNS or physiological saline (control group), probe drugs caffeine (10 mg/kg; CYP1A2 activity), tolbutamide (15 mg/kg; CYP2C9 activity), metoprolol (20 mg/kg; CYP2D6 activity) and dapsone (10 mg/kg; CYP3A4 activity) were administered to rats by intraperitoneal injection. The blood was then collected at different times for ultra performance liquid chromatography/tandem mass spectrometry (UPLC‐MS/MS) analysis. The data showed that PNS exhibited an induction effect on CYP1A2 by decreasing caffeine C_max (36.3%, p < 0.01) and AUC_0‐∞ (22.77%, p < 0.05) and increasing CL/F (27.03%, p < 0.05) compared with those of the control group. Western blot analysis was used to detect the effect of PNS on the protein level of CYP1A2, and the results showed that PNS could upregulate the protein expression of CYP1A2. However, no significant changes in CYP2C9, 2D6 or 3A4 activities were observed. In conclusion, the results indicate that PNS could induce CYP1A2, which may affect the disposition of medicines primarily dependent on the CYP1A2 pathway. Our work may be the basis of related herb–drug interactions in the clinic. Copyright © 2011 John Wiley & Sons, Ltd.     

黄药子与当归配伍对大鼠肝脏CYP1A2、CYP2E1基因mRNA表达的影响

目的：观察单用黄药子和黄药子配伍当归后对大鼠肝CYP1A2、CYP2E1基因mRNA表达的影响。方法：采用逆转录聚合酶链反应（RT-PCR）法测定大鼠肝CYP1A2、CYP2E1基因mRNA在给药后的变化。结果：黄药子组明显诱导CYP1A2、CYP2E1基因mRNA表达，黄药子配伍当归后mRNA表达受到抑制。结论：黄药子诱导P450酶系CYP1A2和CYP2E1mRNA的表达，导致肝中毒。当归配伍黄药子后抑制mRNA表达拮抗肝中毒。     

齐墩果酸对健康人体CYP1A2,CYP2E1及CYP3A4抑制作用的研究

 目的在人体内研究齐墩果酸对CYP1A2,CYP2E1及CYP3A4酶活性的影响,以预测齐墩果酸与常用临床药物的相互作用。方法分别以咖啡因、氯唑沙宗和咪哒唑仑作为CYP1A2,CYP2E1及CYP3A4的探药,采用随机、开放、双周期交叉设计,12名健康男性受试者在服用7d齐墩果酸前后均服用100mg咖啡因、400mg氯唑沙宗和7.5mg咪哒唑仑,服探药后采血测定探药及相应代谢产物的浓度,并计算相关参数。探药和代谢物的浓度分别用RP-HPLC和HPLC-MS测定。结果服用齐墩果酸7d后,咖啡因的代谢受到显著的抑制,其达峰时间、消除半衰期及药-时曲线下面积显著增加;氯唑沙宗的代谢受到轻微抑制,达峰浓度、达峰时间、消除半衰期及药-时曲线下面积均有升高趋势,但无显著性差异;咪哒唑仑的代谢未受影响。结论服用7d齐墩果酸对CYP1A2体内活性有显著抑制作用,对CYP2E1体内活性有轻微抑制作用,而对CYP3A4酶活性无影响。     

清开灵注射液对大鼠CYP1A2和2D6的影响

目的：通过清开灵注射液的大鼠体内、外实验,观察清开灵注射液对大鼠CYP1A2亚型,CYP2D6亚型的影响。方法：通过HPLC法测定全血中咖啡因的代谢率,观测清开灵注射液对大鼠CYP1A2活性的影响；通过HPLC法测定大鼠肝微粒体重组系统非那西丁的代谢比率,确定清开灵注射液对大鼠肝微粒体CYP1A2亚型的作用；测定大鼠肝微粒体重组系统右美沙芬的代谢比率,确定清开灵注射液对大鼠肝微粒体CYP2D6亚型的作用。结果：实验组中给予大鼠不同浓度的清开灵注射液(0.15,0.3,0.6 mL·kg^-1),其咖啡因代谢率为(15.9±3.8)%,(14.5±1.8)%,(12.3±1.2)%,对照组为(16.8±5.9)%,各剂量组及对照组间均无显著性差异；肝微粒体体外重组系统中,实验组各浓度清开灵注射液对CYP2D6没有影响；高剂量组清开灵注射液对CYP1A2有抑制作用。结论：清开灵注射液对CYP1A2和 CYP2D6的活性没有影响。     

一步法分析细胞色素P450 2D6酶活性缺陷等位基因CYP2D6A和CYP2D6B

 目的：CYP2D6A和CYP2D6B是引起细胞色素P4502D6(CYP2D6)酶活性缺陷的最主要的等位基因 ，对CYP2D6A和CYP2D6B的检测可准确(>92%)预测CYP2D6慢代谢者。本研究利用等位基因特异扩增法，建立了一步PCR法测定CYP2D6A和CYP2D6B等位基因。方法：等位基因特异扩增法分析CYP2D6A和CYP2D6B等位基因；右美沙芬作为探针药物测定表型。结果：经130例测定，说明本法更为快捷、更少污染。结论：本法的建立为该项测定应用于临床、指导临床合理用药奠定基础。     

何首乌水提物对大鼠肝脏CYP1A2,CYP2E1酶活性及mRNA表达抑制作用研究

大鼠连续灌胃不同剂量的何首乌水提物(1,10g·kg-1)7 d,取肝脏制备肝微粒体,分别用Cocktail体外孵育法和实时荧光定量PCR技术观察何首乌水提物对大鼠肝脏主要CYP450酶活性及mRNA表达的影响.与空白对照组相比,1,10g·kg-1何首乌水提物组大鼠肝脏CYP2E1酶活性和mRNA的表达均受到明显抑制(CYP2E1酶活性,P＜0.01;CYP2E1的mRNA表达1 g·kg-1组P＜0.05,10 g·kg-1组P＜0.01),CYP3A1酶活性虽显示明显升高(P＜0.01),但mRNA的表达没有显著变化;10 g·kg-1何首乌水提物组大鼠肝脏CYP1A2酶活性和mRNA的表达受到明显抑制(P＜0.01).     

丹参酚酸A对大鼠肝微粒体细胞色素P450酶系的影响

目的:研究丹参酚酸A对大鼠肝微粒体细胞色素P450和细胞色素b_5含量以及CYP1A2和CYP2E1活性的影响.方法:将大鼠分成溶剂对照组和丹参酚酸A给药组,每组10只,雌雄各半,丹参酚酸A给药组尾静脉注射给予丹参酚酸A 20 mg·kg~(-1)·d~(-1),连续给药5 d;溶剂对照组给予相同剂量的溶剂,紫外分光光度法测定大鼠肝微粒体细胞色素P450和细胞色素b_5含量;探针底物法评价CYP1A2和CYP2E1的活性.结果:丹参酚酸A尾静脉注射连续给药5 d后,大鼠细胞色素P450和细胞色素b_5含量与对照组比较均无显著性差异;CYP1A2和CYP2E1的活性与对照组比较也无显著性差异.结论:丹参酚酸A对CYP1A2和CYP2E1没有诱导或抑制作用,与经过CYP1A2和CYP2E1代谢的药物发生相互作用的可能性较小.     

粗叶悬钩子总生物碱对急性肝损伤大鼠肝组织CYP2E1与CYP3A1酶mRNA表达的影响

目的 研究粗叶悬钩子总生物碱对肝药物代谢酶CYP2E1和CYP3A1 mRNA表达的影响。     

Cocktail法评价注射用益气复脉(冻干)对大鼠肝微粒体CYP1A2、CYP3A的诱导作用

目的:研究注射用益气复脉(冻干)Yi Qi Fu Mai Injection(YQFM)(Lyophilized)对大鼠肝微粒体CYP1A2、CYP3A的诱导作用。方法:将wistar大鼠分为生理盐水对照组,苯巴比妥钠诱导组,YQFM组,连续给药7天后处死,制备肝微粒体。采用Cocktail法,将特异性探针底物非那西丁(CYP1A2)、睾酮(CYP3A)与肝微粒体共孵育,采用高效液相色谱测定孵育所得代谢产物对乙酰氨基酚和6β-羟基睾酮的生成速率,来评价YQFM对CYP1A2、CYP3A的诱导作用。结果:空白对照组、诱导组和YQFM组的对乙酰氨基酚的生成速率分别为(18.04±1.00)、(43.07±2.90)、(27.6±4.5)ng.(mg pro-tein)-1.min-1,6β-羟基睾酮的生成速率分别为(15.79±1.43)、(40.86±3.32)、(32.8±3.67)ng.(mg protein)-1.min-1。结论:YQFM对大鼠肝微粒体CYP1A2、CYP3A有诱导作用。     

葛根散对急性酒精性肝损伤小鼠肝微粒体 CYP450含量及CYP2E1活性的影响

目的:考察经典解酒方葛根散对急性酒精性肝损伤小鼠肝微粒体细胞色素P450( CYP450)含量及细胞色素P450-2E1( CYP2E1)活性的影响.方法:实验小鼠分葛根散高、中、低剂量组、正常对照组和模型对照组,各给药组分别给予葛根散水煎剂(按生药量计,20,10,5 g·kg-1)ig,对照组代以等体积蒸馏水,每天1次,连续10 d;每天给药30 min后,模型对照组和各给药组以15 mL·kg-1 56％白酒ig,正常对照组代以等体积蒸馏水,以血清丙氨酸转氨酶(ALT)、天冬氨酸转氨酶(AST)及肝脏系数为参考指标,制备急性酒精性肝损伤小鼠模型;取小鼠肝组织检测葛根散对肝微粒体CYP450含量及CYP2E1活性的影响.结果:与正常对照组比较:模型对照组血清ALT、AST水平及肝脏系数显著升高(P＜0.01),肝微粒体蛋白含量无明显变化,CYP450含量及CYP2E1活性显著提高(P＜0.01).与模型对照组比较:葛根散各剂量组血清ALT,AST水平及大剂量组肝脏系数呈明显降低(P ＜0.05或P＜0.01),肝微粒体蛋白含量无明显变化,CYP450含量明显增多(P＜0.01)且有剂量差异性(P＜0.05),CYP2E1活性显著降低(P＜0.01)且有剂量差异性(P＜0.01).结论:葛根散对急性酒精性肝损伤小鼠肝功能具有良性干预作用,能提高酒精性肝损伤小鼠肝微粒体CYP450含量和降低CYP2E1活性.     

桂枝汤对大鼠肝微粒体细胞色素P450酶活性的影响

目的:研究桂枝汤对大鼠肝微粒体细胞色素P450酶CYP1A2、CYP2D6和CYP3A4活性的影响。方法:18只大鼠被随机分成3组,每组6只。以生理盐水为空白对照,大鼠每日灌胃给予桂枝汤10 g/kg,连续7 d,测定其肝微粒体CYP1A2、CYP2D6和CYP3A4活性。结果:与对照组相比,桂枝汤对大鼠CYP1A2、CYP2D6和CYP3A4的活性无明显差异(P>0.05)。结论:桂枝汤对大鼠CYP1A2、CYP2D6和CYP3A4活性无影响。     

Andrographolide inhibits the expression and metabolic activity of cytochrome P450 3A4 in the modified Caco-2 cells

Aim of the study

The aim of this study is to examine the effects of andrographolide on intestinal enzyme cytochrome P450 3A4 (CYP3A4) and predict whether oral administration of andrographolide-containing remedy leads to herb–drug interaction.

Materials and methods

Caco-2 cells are treated with 1α, 25-dihydroxyvitamin D3 for 3 wks to induce the expression of CYP3A4, and then andrographolide (1, 10, 100 μM) is added and treated for 72 h. Upon the further 4-h testosterone (250 μM) or nifedipine (200 μM) treatment, the basolateral medium samples and the Caco-2 monolayers are collected for analyses.

Results

Andrographolide (1, 10, 100 μM) significantly down-regulates the mRNA level and protein level of CYP3A4, and inhibits nifedipine oxidation and testosterone 6β-hydroxylation.

Conclusion

Oral administration of andrographolide likely leads to reduction of the metabolic activity of intestinal CYP3A4, therefore herb preparations containing andrographolide may result to herb–drug interactions in combination therapy.     

血塞通注射液等36种中药对大鼠CYP2D1的作用

目的:研究血塞通注射液等36种中药对大鼠肝微粒体CYP2D1的作用。方法:建立右美沙芬的高效液相-紫外测定方法;以右美沙芬为探针药物,通过考察其体外转化率的变化评价受试中药对大鼠CYP2D1的作用。结果:血塞通注射液、清开灵注射液、银黄口服液、蓝芩口服液使探针药物的转化率分别为(218.7±11.6)pmol/min.mg、(214.9±19.7)pmol/min.mg、(204.3±13.3)pmol/min.mg、(127.0±41.4)pmol/min.mg,显著低于对照组(431.5±4.2)pmol/min.mg,四种药物的IC50分别为1.0ml/100ml、0.6ml/100ml、0.2ml/100ml、0.1ml/100ml。结论:血塞通注射液、清开灵注射液、银黄口服液、蓝芩口服液体外对大鼠CYP2D1有显著抑制作用,且呈浓度依赖性。