芍药苷微乳溶液在大鼠在体肠的吸收动力学研究

目的考察芍药苷微乳溶液在大鼠在体肠吸收动力学特征,并与其水溶液进行比较,研究芍药苷微乳溶液在各肠段的吸收情况。方法采用大鼠在体肠吸收实验方法,用HPLC和UV法分别测定循环液芍药苷和酚红的浓度。结果在2～20mg.L-1浓度范围内芍药苷微乳溶液的吸收量与浓度成线性关系,吸收速率常数(Ka)值基本保持不变,芍药苷微乳溶液吸收药量和Ka均高于其水溶液;各肠段吸收有差异,结肠吸收药量和Ka明显高于十二指肠、空肠和回肠,Ka值依次为(0.501±0.031)、(0.086±0.003)、(0.108±0.017)、(0.114±0.006)h-1。结论芍药苷微乳溶液在肠道的吸收呈一级吸收动力学,吸收机制为被动吸收;芍药苷微乳溶液在空肠和结肠段吸收较好,且在肠道吸收好于芍药苷水溶液。提示可将芍药苷制成微乳制剂,以提高其生物利用度。     

芍药内酯苷抗抑郁作用的实验研究

目的:研究芍药内酯苷的抗抑郁作用.方法:采用小鼠悬尾、小鼠强迫游泳、小鼠自主活动实验,观察芍药内酯苷对小鼠游泳不动时间、悬尾不动时间及自主活动的影响.结果:芍药内酯苷14、7mg/kg均可显著缩短小鼠悬尾( P＜0.01,P＜0.05)及强迫游泳不动时间(P＜0.05),实验各剂量芍药内酯苷对小鼠自主活动无明显影响.结论:芍药内酯苷具有明显的抗抑郁作用.     

RP—HPLC法同时测定白芍中芍药苷和芍药内酯苷的含量

目的:建立了RP-HPLC法对白芍中芍药苷和芍药内酯苷同时定量,考察不同产地白芍中芍药苷、芍药内酯苷的含量。方法:高效液相色谱法,Hypersil-C_(18)色谱柱(4.6 mm×200 mm,5 μm),流动相:甲醇-乙腈-水(10:10:80),流速0.8 mL·min~(-1),检测波长230nm,柱温为室温。以咖啡因为内标测定了白芍中芍药苷和芍药内酯苷的含量。结果:芍药苷、芍药内酯苷浓度与峰面积呈良好的线性关系,线性范围分别是:芍药苷18.24～273.6μg·mL~(-1),r=0.999 6(n=7);芍药内酯苷4.96～74.4μg·mL~(-1),r=0.999 7(n=7);回收率(n=9)芍药苷为97.5％～97.7％,芍药内酯苷为91.1％～96.3％。结论:本方法测定了26个不同产地、不同批号及炮制品的白芍样品中芍药苷、芍药内酯苷含量,该方法分离度好,快速,简便,重现性好。     

不同比例配伍的赤芍-藜芦药对中芍药苷和芍药苷内酯的含量测定

目的考察不同比例赤芍与藜芦配伍后芍药苷和芍药苷内酯的含量变化。方法采用HPLC法测定芍药苷和芍药苷内酯的含量;色谱条件:Inertsil C_(18)柱(250 mm×4.6 mm,5μm);流动相为乙腈-0.1 mL·L~(-1)磷酸水溶液(13∶87);检测波长:230 nm;流速:1.0 mL·min~(-1)。结果芍药苷和芍药苷内酯质量浓度分别在7.03～450.00和1.13～290.00μg·mL~(-1)(r=1.000 0)范围内与峰面积呈良好线性关系,平均回收率分别为97.93%(RSD=1.85%)和97.15%(RSD=1.81%)。结论赤芍与藜芦不同比例配伍后芍药苷和芍药苷内酯含量均比赤芍单煎液低,存在减效现象,赤芍-藜芦药对可能为相反配伍。     

HPLC-MS/MS法同时测定坤复康片中5种有效成分的含量

目的采用HPLC-MS/MS法同时测定坤复康片中氧化苦参碱、苦参碱、芍药内酯苷、芍药苷及苯甲酰芍药苷等5种有效成分的含量。方法待测样品研磨后采用甲醇超声提取,以Phenomenex Luna C18(150 mm×4.6 mm,5μm)为色谱柱,采用体积分数为0.1%的甲酸乙腈溶液(A)-体积分数为0.1%的甲酸溶液(B)为流动相,梯度洗脱,流速为0.3 m L·min-1,分析时间为9.0 min。质谱ESI源-MRM模式,正、负离子同时检测。5种成分的监测离子对分别为:265.2/247.4(氧化苦参碱)、249.3/148.4(苦参碱)、479.3/197.1(芍药内酯苷)、525.1/121.0(芍药苷)、583.3/120.8(苯甲酰芍药苷)。建立的方法对3批坤复康片进行了测定。结果氧化苦参碱、苦参碱、芍药内酯苷、芍药苷及苯甲酰芍药苷5种有效成分在测定浓度范围内均具有良好的线性关系,r>0.994 8,回收率为95.8%~102.1%。结论用本方法可以同时测定5种有效成分,适用于同时测定坤复康片中氧化苦参碱、苦参碱、芍药内酯苷、芍药苷及苯甲酰芍药苷的含量,同时能较好地控制坤复康片的质量。     

硫磺熏制白芍的安全性评价初步研究

建立硫磺熏制白芍中芍药苷及其衍生物芍药苷亚硫酸酯、芍药内酯苷的高效液相色谱含量测定方法,并通过对芍药苷亚硫酸酯的细胞毒性以及白芍药材的小鼠急性毒性评价,对硫磺熏制前后的白芍进行安全性评价研究。含量测定采用Shiseido Capcell PAK C18(250 mm×4.6 mm ID,5μm)色谱柱,流动相乙腈—0.02%磷酸水溶液(15∶85),检测波长230 nm,流速1.0 mL.min-1,柱温30℃;采用MTT法考察芍药苷亚硫酸酯对小鼠原代肝细胞和人原代肝细胞的毒性;采用小鼠灌胃最大给药量法考察芍药苷亚硫酸酯和白芍的急性毒性。芍药苷亚硫酸酯、芍药苷和芍药内酯苷分别在0.041 8～1.045 0 mg.mL-1、0.023 5～0.587 5 mg.mL-1和0.039 8～0.995 0 mg.mL-1内呈良好线性,r>0.999 8,平均回收率99.11%～101.71%,RSD<2%;芍药苷亚硫酸酯的浓度≤300μmol.L-1,各检测浓度均无明显细胞毒性;芍药苷亚硫酸酯水溶液和白芍水提物分别进行小鼠灌胃给药,最大耐受量分别为5 g.kg—1和80 g.kg-1。建立的白芍中3个成分含量测定方法简单快速、准确可靠、重复性好;硫磺熏制后白芍中芍药苷的含量显著降低,且芍药苷亚硫酸酯的含量与芍药苷的含量呈负相关;体外肝细胞和体内小鼠急性毒性实验未发现芍药苷亚硫酸酯的毒性,硫磺熏制白芍及白芍水提物比较无显著的毒性。     

赤芍与辛芍组方中没食子酸、芍药内酯苷在大鼠体内的药动学比较

目的建立UPLC-MS法,进行辛芍组方与单味赤芍提取物没食子酸、芍药内酯苷大鼠药动学研究,探讨中药复方对药效成分体内过程的影响。方法色谱采用Waters BEH C18柱,0.1%甲酸乙腈-0.1%甲酸水梯度洗脱,质谱采用电喷雾电离源(ESI),扫描方式为选择性离子监测(SIR)。实验大鼠分别灌胃辛芍组方、赤芍提取物,测定不同时间点大鼠血浆没食子酸和芍药内酯苷浓度,采用DAS2.0药动学软件对参数进行拟合。结果没食子酸和芍药内酯苷线性关系、精密度、准确度和稳定性良好。没食子酸、芍药内酯苷在大鼠体内符合二室药动学模型,赤芍和辛芍组方两组没食子酸T max和T1/2z差异无显著性,芍药内酯苷的各药动学参数均具有一定差异。结论中药组方与单味药材体内药动学之间存在差异,其产生的原因可能与复方间的相互配伍以及复方组成有关。     

高效液相色谱法测定白芍药材中芍药苷和芍药内酯苷的含量

目的:建立高效液相色谱法测定白芍药材中芍药苷和芍药内酯苷含量的方法.方法:采用高效液相色谱法,色谱柱:Kromasil ODS2 (4.6 mm×250 mm,5 μm),流动相:甲醇-0.1％磷酸溶液(用1％氢氧化钠溶液调节pH值至4.2)(34:66);流速:1.0 mL·min-1,检测波长:230 nm.结果:芍药内酯苷在0.018～0.576 μg范围内呈线性关系(r=0.9999),芍药苷在0.028～0.896μg范围内呈线性关系(r=0.9999);芍药内酯苷加样回收率为98.18％(n=6),RSD值小于2％,芍药苷加样回收率为98.26％(n=6),RSD值小于2％.结论:本研究所建方法灵敏、简便,能同时准确测定白芍药材中芍药苷和芍药内酯苷的含量.     

单向肠灌流法研究芍药苷和刺芒柄花苷的肠吸收特性

目的:采用在体单向肠灌流法研究芍药苷和刺芒柄花苷在大鼠不同肠段的吸收特性,探索复方给药对单一活性成分吸收的影响。方法:采用高效液相色谱法进行含量测定和方法学验证。色谱柱:Shim Pack ODS C₁₈(250 mm×4.6 mm,5μm);流动相:测定芍药苷时为甲醇-乙腈-1.0 mL·L^-1磷酸水溶液(15∶15∶70),测定刺芒柄花苷时为乙腈-1.0 mL·L^-1甲酸水溶液(55∶45);流速:1.0 mL·min^-1;检测波长:230 nm(芍药苷)、254 nm(刺芒柄花苷);柱温:30℃。通过对相同药物浓度在不同肠段、不同药物浓度对同一肠段有效渗透系数(Peff)的影响研究芍药苷和刺芒柄花苷的主要吸收途径,在灌流液中添加维拉帕米研究糖蛋白抑制剂对药物在十二指肠Peff的影响,通过比较单一成分与复方灌流后的Peff差异,研究复方形式对单一活性成分在肠道吸收的影响。结果:芍药苷和刺芒柄花苷的质量浓度均在0.5~60μg·mL^-1范围内,与相应的峰面积呈良好的线性关系,相关系数分别为0.9999和0.9998;日内精密度RSD分别小于2.5%和2.9%,日间精密度RSD均小于3.1%,稳定性RSD分别小于3.6%和3.3%。芍药苷和刺芒柄花苷在大鼠小肠的Peff由大到小顺序均为十二指肠、空肠、回肠。糖蛋白抑制剂维拉帕米对二者在十二指肠的吸收无显著影响,同时不同浓度的含药灌流液的Peff又具有显著的差别,具有浓度饱和现象,复方灌流液补阳还五汤中芍药苷和刺芒柄花苷的Peff高于单一成分灌流。结论:十二指肠为芍药苷和刺芒柄花苷最佳吸收肠段,以复方形式给药能够提高部分有效成分在肠道的吸收。     

芍药内酯苷对高糖诱导的人脐静脉内皮细胞损伤的保护作用

目的 研究芍药内酯苷对高糖损伤的人脐静脉内皮细胞（HUVECs）的保护作用及机制.方法 采用33mmol·L-1的高糖培养基建立HUVECs损伤模型,并在造模前给予不同浓度的芍药内酯苷进行预保护,采用MTT法检测细胞活力,流式细胞术检测细胞凋亡,采用试剂盒检测细胞中内源性一氧化氮合酶（eNOS）和半胱天冬酶-3（Caspase-3）活性以及培养液中一氧化氮（NO）和乳酸脱氢酶（LDH）含量.结果 芍药内酯苷能够剂量依赖性增强细胞活力,降低Caspase-3活性和细胞凋亡率（P＜0.05）,并能增强eNOS的活性和NO的释放量（P＜0.05）.结论 芍药内酯苷对高糖诱导的HUVEs损伤具有保护作用,其机制可能与促进eNOS活性提高、NO释放量增加和抑制Caspase-3活性有关.     

芙朴感冒颗粒中橙皮苷小肠吸收特征研究

目的 研究芙朴感冒颗粒中的橙皮苷跨膜吸收特征及其影响因素.方法 使用Caco-2细胞模型考察橙皮苷的小肠吸收,研究不同药物浓度、pH值、温度和抑制剂对橙皮苷在transwell细胞培养板中从顶膜(AP)到基底(BL)的双向渗透吸收的影响.采用药动学实验研究芙朴感冒颗粒在体内对橙皮苷吸收的影响.结果 橙皮苷以3种测试浓度...     

抗肿瘤药物西美替尼在Caco-2细胞模型中的吸收转运

目的研究抗肿瘤药物西美替尼在Caco-2细胞模型中的吸收转运。方法建立Caco-2细胞转运模型,采用HPLC法测定药物浓度,计算表观渗透系数(P_app),研究西美替尼的Caco-2细胞跨膜转运情况。结果西美替尼在转运过程中没有明显的浓度依赖性。在低浓度范围内,药物的转运速率随着浓度的增加而增加;在较高浓度时达到饱和。西美替尼不同浓度的P_app值(3.91×10^-5、3.29×10^-5、1.90×10^-5和0.95×10^-5cm·s^-1)基本都大于难吸收药物的临界值(1×10^-5cm·s^-1)。西美替尼在Caco-2细胞中的转运呈现较强的方向性,从肠腔面到基底面的P_app值显著大于从基底面到肠腔面的P_app值（2.36～7.58倍）。ATP抑制剂叠氮化钠能显著降低西美替尼的正向转运程度,并升高其反向转运程度。当加入葡萄糖后,从肠腔面到基底面的跨膜转运程度显著降低。结论西美替尼主要是以转运载体介导的主动转运方式被吸收,其小肠吸收情况较好。     

The blood-brain barrier permeability of 20(S) and 20(R)-protopanaxatriol epimers and dammar-20(22)E,24-diene-3β,6α,12β-triol in MDCK-pHaMDR cell monolayer model

The blood-brain barrier permeability of 20(S) and 20(R)-protopanaxatriol epimers and dammar-20(22)E,24-diene-3β,6α,12β-triol were investigated using the MDCK-pHaMDR cell monolayer model. The bidirectional permeability tests were carried out, and the apparent permeability coefficients (P_app) were calculated. The two protopanaxatriol epimers showed good permeability with P_app values of ~10^–5 cm/s, whereas dammar-20(22)E,24-diene-3β,6α,12β-triol showed poor permeability with P_app of <1×10^–7 cm/s. The three compounds showed differences in intracellular accumulations due to their different structures. Inhibition of P-gp with verapamil showed that the transport mechanisms in MDCK-pHaMDR cell monolayer for compounds 1and 2 epimers were not only simple passive diffusion but also involving an efflux way mediated by P-gp. These findings provided new basis for the further study of compounds 1 and 2 acting on the brain.     

DAD-HPLC法同时测定戊己丸中4种成分的含量

目的:建立戊己丸中芍药苷、盐酸小檗碱、吴茱萸碱与吴茱萸次碱4种成分的含量测定方法。方法:采用高效液相色谱法,应用XSelect CSH-C₁₈(4.6 mm×150 mm,5μm)色谱柱,以乙腈-0.1%磷酸溶液(含0.05 mol·L^-1磷酸二氢钾)为流动相,梯度洗脱,流速为1.0 mL·min^-1,柱温30℃,检测波长:芍药苷232 nm,盐酸小檗碱230 nm,吴茱萸碱226 nm,吴茱萸次碱342 nm。结果:芍药苷、盐酸小檗碱、吴茱萸碱、吴茱萸次碱分别在3.856×10²~3.856×10³ ng、4.239×10²~4.239×10³ ng、16.46~1.646×10² ng、8.340~83.40 ng范围内与峰面积呈良好的线性关系,加样回收率分别为98.7%、103%、95.3%、94.3%。结论:本方法专属性强,前处理操作简单,检测结果准确,可为戊己丸全面质量控制提供参考。     

补阳还五汤抗动脉粥样硬化8个效应成分UPLC-MS/MS测定方法研究

目的:建立补阳还五汤抗动脉粥样硬化8个效应成分(羟基红花黄色素A、芍药内酯苷、芍药苷、苦杏仁苷、黄芪甲苷、正丁烯基苯酞、Z-蒿本内酯和阿魏酸)的UPLC-MS/MS含量测定分析方法。方法:基于Citespace软件进行文献分析,明确补阳还五汤中8个抗动脉粥样硬化效应成分。采用ACQUITY UPLC BEH C₁₈(50 mm×2.1 mm,1.7μm)色谱柱,柱温40℃,以0.1%甲酸水(A)-乙腈(B)为流动相,梯度洗脱,进样量0.3 mL·min^-1,电喷雾离子化(ESI)源,正离子模式扫描,多反应监测模式(MRM)检测各有效成分。结果:羟基红花黄色素A的质量浓度在10.00~1350.00 ng·m L^-1的范围内线性关系良好(r=0.9999),芍药内酯苷、芍药苷和苦杏仁苷的质量浓度均在2.00~2700.00 ng·m L^-1的范围内线性关系良好(r=0.9999),黄芪甲苷、正丁烯基苯酞和Z-蒿本内酯的质量浓度均在2.00~270.00 ng·mL^-1的范围内线性关系良好(r=0.9999),阿魏酸的质量浓度在5.00~675.00 ng·mL^-1的范围内线性关系良好(r=0.9998);平均回收率分别为94.6%、100.5%、96.3%、91.8%、108.2%、104.1%、99.8%、105.2%,RSD分别为8.5%、9.0%、5.2%、3.7%、3.9%、5.0%、7.1%、6.2%。补阳还五汤样品中上述抗动脉粥样硬化8个效应成分含量的RSD均<5%。结论:该方法重复性、回收率好,可为抗动脉粥样硬化复方补阳还五汤质量控制提供分析方法。     

Uptake and transport of the ACE-inhibitor ceronapril (SQ 29852) by monolayers of human intestinal absorptive (Caco-2) cells in vitro

The angiotensin-converting enzyme (ACE)-inhibitor ceronapril (SQ 29852) is shown to be a substrate of the intestinal dipeptide transporter. Uptake by Caco-2 cells, grown as confluent monolayers, follows a major saturable pathway (K_m, 0.91 ± 0.11 mM; 90% at 1 mM) together with a minor passive component (k_J, 32.3 ± 6.6 ng (10⁶ cells)⁻¹ (20 min)⁻¹. Uptake was inhibited by competition with dipeptides such as l-AIa-l-Pro (K_i, 2.96 mM) and l-Phe-Gly (K_i, 3.84 mM) but not by cephalosporins such as cephalexin. In contrast, transport was non-saturable, flux increased linearly with concentration and data were consistent with a passive transepithelial transport mechanism. Transport profiles showed a biphasic dependence upon time with an initial flux of 0.83 ± 0.02 ng insert⁻¹ min⁻¹ (k₁) and a terminal value of 1.65 ± 0.08 ng insert⁻¹ min⁻¹ ((k₂) at 100 μM. It is concluded that the basolateral efflux is retarded so that the passive paracellular transport controls the overall transepithelial transport characteristics in the Caco-2 model. Carrier-mediated uptake into intestinal enterocytres, followed by rate-limiting basolateral efflux, may explain the extended t_max in vivo following oral administration.     

Drug permeability in 16HBE14o- airway cell layers correlates with absorption from the isolated perfused rat lung

The permeability of the lung is critical in determining the disposition of inhaled drugs and the respiratory epithelium provides the main physical barrier to drug absorption. The 16HBE14o- human bronchial epithelial cell line has been developed recently as a model of the airway epithelium. In this study, the transport of 10 low molecular weight compounds was measured in the 16HBE14o- cell layers, with apical to basolateral (absorptive) apparent permeability coefficients (P_app) ranging from 0.4 × 10⁻⁶ cm s⁻¹ for Tyr-D-Arg-Phe-Phe-NH₂ to 25.2 × 10⁻⁶ cm s⁻¹ for metoprolol. Permeability in 16HBE14o- cells was found to correlate with previously reported P_app in Caco-2 cells and absorption rates in the isolated perfused rat lung (k_a,lung) and the rat lung in vivo (k_{a,in vivo}) [Tronde, et al., 2003. J. Pharm. Sci. 92, 1216–1233; Tronde, et al., 2003. J. Drug Target. 11, 61–74]. Log linear relationships were established between P_app in 16HBE14o- cells and P_app in Caco-2 cells (r² = 0.82), k_a,lung (r² = 0.78) and k_{a,in vivo} (r² = 0.68). The findings suggest that permeability in 16HBE14o- cells may be useful to predict the permeability of compounds in the lung, although no advantage of using the organ-specific cell line 16HBE14o- compared to Caco-2 cells was found in this study.     

Mechanisms Responsible for Poor Oral Bioavailability of Paeoniflorin: Role of Intestinal Disposition and Interactions with Sinomenine

Purpose To determine the intestinal disposition mechanisms of paeoniflorin, a bioactive glucoside, and to investigate the mechanisms by which sinomenine increases paeoniflorin bioavailability. Materials and Methods A single-pass “four-site” rat intestinal perfusion model and a cultured Caco-2 cell model were employed. Results In both model systems, paeoniflorin permeability was poor. In the perfusion model, maximal absorption and metabolism of paeoniflorin occurred in duodenum and jejunum, which were significantly decreased by a glucosidase inhibitor gluconolactone (20 mM). On the other hand, paeoniflorin absorption in terminal ileum increased significantly but its metabolism did not in the presence of sinomenine and cyclosporine A. In the Caco-2 cell model, paeoniflorin was transported 48-fold slower than its aglycone (paeoniflorigenin). Absorptive transport of paeoniflorin was significantly (p < 0.05) increased by sinomenine (38%), verapamil (27%), and cyclosporine A (41%), whereas its secretory transport was significantly (p < 0.01) decreased by sinomenine (50%), verapamil (35%) and cyclosporine A (37%). In contrast, MRP inhibitors MK-571 and leukotriene C4 did not affect transport of paeoniflorin. Lastly, sinomenine was also shown to significantly increase the absorptive transport of digoxin (a prototypical p-glycoprotein substrate) and to significantly decrease its secretory transport. Conclusions Poor permeation, p-gp-mediated efflux, and hydrolysis via a glucosidase contributed to the poor bioavailability of paeoniflorin. Sinomenine (an inhibitor of the p-gp-mediated digoxin efflux) increased paeoniflorin's bioavailability via the inhibition of p-gp-mediated paeoniflorin efflux in the intestine.     

Drug specificity and intestinal membrane localization of human organic cation transporters (OCT)

This study was performed to investigate which human organic cation transporter, hOCT1, hOCT2 or hOCT3, participates with regard to cation specificity and membrane localization in the intestinal absorption of orally available cationic drugs. Inhibition of N-[methyl-³H]4-phenylpyridinium ([³H]MPP⁺) uptake by various compounds into Caco-2 cells and into cells (HEK-293 or CHO) that were stably transfected with hOCT1, hOCT2 or hOCT3 was compared. The uptake of [³H]MPP⁺ into Caco-2 cells was inhibited by atropine, butylscopolamine, clonidine, diphenhydramine, etilefrine, quinine and ranitidine with IC₅₀ values between 6 μM and 4 mM. Transepithelial, apical to basal flux of [³H]MPP⁺ across Caco-2 cell monolayers was also strongly inhibited by these compounds. The inhibitory potency of the cationic drugs and prototypical organic cations at Caco-2 cells correlated well with the inhibitory potency measured at CHO-hOCT3 cells but much less with that at HEK-hOCT1 and -hOCT2 cells. This is functional evidence for the predominant role of hOCT3. Etilefrine and atropine were specifically transported into CHO cells by hOCT3. In Caco-2 cells, the mRNA of all three hOCT and the proteins hOCT2 and hOCT3 were detected. More importantly, immunocytochemical analyses of human jejunum revealed for the first time that hOCT3 is localized to the brush border membrane whereas hOCT1 immunolabeling was mainly observed at the lateral membranes of the enterocytes.