Pretreatment with potent P-glycoprotein ligands may increase intestinal secretion in rats

The expression of P-glycoprotein is induced in cell cultures upon exposure to various inducers. Therefore, the aim of the present study was to evaluate the in-vivo relevance of this observation, i.e. the influence of chronic pretreatments with selected drugs -- all of which are ligands to P-glycoprotein (P-gp) as demonstrated in radioligand binding studies and all of which have some or a considerable effect on P-gp expression in Caco-2 cells -- on the effective intestinal permeabilities of the model compound talinolol in rats employing in-situ single-pass intestinal perfusion of three different gut segments. Talinolol was selected, because it shows high selectivity for one of the exsorptive transporters (P-gp) and its intestinal permeability is very sensitive to changes in exsorption when the perfusate concentration is low. Prior to the induction study the perfusion model was optimized regarding the type and concentration of a competitive inhibitor which may be used to block the exsorption-related permeability reduction (through intestinal exsorption) during an ongoing perfusion and would permit an intra-individual comparison of the effective permeability without and with blockade of exsorption. While repetitive verapamil and talinolol dosing had no statistically significant exsorption-inducing effect, vinblastine and rifampicin pretreatments resulted in decreased intestinal talinolol permeabilities in the three tested gut segments, duodenum, jejunum, and colon [e.g., S-talinolol in jejunum: control, 2.50 x 10(-4) cm/s; vinblastine induction, 1.48 x 10(-4) cm/s (P<0.05); rifampicin induction, 1.51 x 10(-4) cm/s (P<0.05)]. Addition of an efficient secretion inhibitor (vinblastine) to the perfusate permitted the determination of the impact of inhibitable secretory processes on the total effective permeabilities and an estimation of passive permeability in the respective individual. The inhibitable permeability fractions were higher for vinblastine than for any other pretreatment and the difference from control pretreatment was statistically significant for all intestinal segments (duodenum, 61.8%; jejunum, 63.1%; colon, 43,7%; S-talinolol). Statistically significant differences were also detected for rifampicin in the perfused duodenum and jejunum (33.1 and 27.5% increase in inhibitable fraction, respectively, for S-talinolol). These differences are explained by a significant induction of outside-directed transport in the intestinal enterocytes by vinblastine and rifampicin.     

环糊精包合作用对在体大鼠肠道P-糖蛋白药泵的影响

目的研究环糊精包合作用对P-糖蛋白底物药物体内吸收的影响。方法以P-糖蛋白底物盐酸小檗碱为模型药物,使用大鼠在体单向肠灌流装置,采用高效液相色谱法分别测定灌流液中盐酸小檗碱和酚红的浓度变化;研究2-羟丙基-β-环糊精(HP-β-CyD)对盐酸小檗碱肠道吸收的影响,以此评价环糊精包合作用对P-糖蛋白药泵的影响。结果盐酸小檗碱、盐酸小檗碱/HP-β-CyD物理混合物及盐酸小檗碱/HP-β-CyD包合物在大鼠空肠的吸收速率常数(Ka)分别为(0.45±0.029)、(0.70±0.087)、(2.39±0.119)×10-2·min-1,有效渗透系数(Peff)分别为(0.43±0.028)、(0.63±0.098)、(2.17±0.145)×10-3min·cm-1。HP-β-CyD对盐酸小檗碱的包合作用促进了大鼠肠道对盐酸小檗碱吸收。HP-β-CyD与盐酸小檗碱混合给药也能促进药物的吸收,但其作用远低于包合作用。结论环糊精的包合作用将有可能成为提高P-糖蛋白底物药物生物利用度的有效手段。     

Effect of hydroxyzine on the transport of etoposide in rat small intestine

Etoposide, an anti-neoplastic agent and a substrate of P-glycoprotein (P-gp), exhibits variable oral bioavailability. P-gp, the multidrug resistance gene (mdr1) product, has been considered as an absorption barrier against intestinal drug absorption. Terfenadine, an antihistamine, has been shown to be a P-gp inhibitor. The current study was designed to assess the effect of hydroxyzine, an antihistamine, on the transport of etoposide in the small intestine. Everted rat gut sacs were used to determine the absorption and exsorption of etoposide under different conditions, as rhodamine 123 was chosen to evaluate the role of P-gp in the drug interaction. The results showed that the transport of etoposide was significantly increased from the luminal site to the serosal site in the jejunum by 2- and 4-fold after 90 min in the presence of hydroxyzine and quinidine, respectively. A similar trend was observed in the ileal sacs. This in vitro exsorption study also demonstrated that hydroxyzine could reduce the efflux of etoposide to the luminal site in either jejunum or ileum. The effect of hydroxyzine on the pharmacokinetics of etoposide differed by the in vivo route of administration, thus assuming clinical importance for chemotherapeutic treatment.     

Transport of phenobarbitone into the intestinal lumen and the biliary tract following i.v. administration to rats

The exsorption of intravenously administered phenobarbitone into the lumen of the small intestine of rats has been examined by an in-situ single-pass perfusion technique. The levels of the drug in the bile were about twice that in serum. Moreover, it was shown that the drug was appreciably exsorbed into the intestinal lumen. The exsorption rates of the drug into the perfusates were not very different between isotonic phosphate buffers at pH 6.0 and 8.0. The amounts of phenobarbitone exsorbed into the perfusate and excreted into the bile were 6.47 and 0.45% of the dose for isotonic phosphate buffer at pH 6.0, and were 6.09 and 0.54% of the dose for isotonic phosphate buffer at pH 8.0.     

硫酸沙丁胺醇大鼠在体肠吸动力学研究

目的研究硫酸沙丁胺醇在大鼠各肠段的吸收动力学特征。方法采用大鼠在体肠回流法进行动力学试验,从吸收部位、药物浓度、pH值等方面对药物在体内的各个肠段的吸收特性进行研究。结果硫酸沙丁胺醇在大鼠肠道中的吸收不受药物浓度、回流介质pH值等的影响,在分肠段试验中,吸收速率常数Ka(1/h)依次为十二指肠0.052,空肠0.046,回肠0.042,结肠0.030,结肠的吸收显著低于十二指肠、空肠和回肠段;在pH5.4~7.8回流介质中吸收无显著差异,在50~200μg/mL浓度范围内,药物吸收量与浓度呈线性关系。结论硫酸沙丁胺醇在大鼠体内各肠段均有吸收,吸收机制以被动扩散为主,适于制成Tlag<5 h的口服迟释制剂。     

Absorption of benzoic acid in segmental regions of the vascularly perfused rat small intestine preparation.

Oral bioavailability is a consequence of intestinal absorption, exsorption, and metabolism and is further modulated by the difference in activities among segmental regions. The influence of these factors on the net absorption of benzoic acid (BA), a substrate that is metabolized to hippurate and is transported by the monocarboxylic acid transporter 1, was studied in the recirculating, vascularly perfused, rat small intestine preparation. Metabolism of BA was not observed for both systemic and intraluminal injections into segments of varying lengths. But, secretion of BA into lumen was noted. Absorption of BA (0.166-3.68 micromol) introduced at the duodenal end for absorption by the entire intestine was complete (>95% dose at 2 h) and dose-independent, yielding similar absorption rate constants (k(a) of 0.0464 min(-1)). The extent of absorption remained high (92-96% dose) when BA was injected into closed segments of shorter lengths (12 or 20 cm), suggesting a large reserve length of the rat intestine. However, k(a) was higher for the jejunum (0.0519 and 0.0564 min(-1), respectively, for the 12- and 20-cm segments) and exceeded that for the duodenum (12-cm segment, 0.0442 min(-1)) and ileum (20-cm segment, 0.0380 min(-1)) at closed injection sites. The finding paralleled the distribution of monocarboxylic acid transporter isoform 1 detected by Western blotting along the length of the small intestine. Fits of the systemic and oral data (based on duodenal injection for absorption by the whole intestine) to the traditional, physiological model and to the segregated flow model (SFM) that describes partial intestinal flow to the enterocyte region showed a better fit with the SFM even though metabolite data were absent.     

Effect of theophylline on the intestinal clearance of drugs in rats.

Intestinal exsorption of salicylic acid, urea and quinidine was measured during the perfusion of the rat intestinal lumen with Tyrode solution. The intestinal clearance (CLi) of the three compounds was measured by dividing the rate of appearance in the intestinal luminal perfusate by the plasma concentration of the compound. Co-administration of theophylline (0.2 mg h-1) with the test agents increased the CLi of salicylic acid, did not alter the CLi of urea, but decreased the CLi of quinidine. The effect of theophylline on the CLi of quinidine was enhanced with increasing dose. Theophylline was found to increase microclimate-pH at the intestinal surface, but the magnitude of delta pH alone could not explain the effect of theophylline on the CLi of quinidine. The data, together with previous observations, suggest that the intestinal exsorption of drugs was affected by the microclimate pH and by the unstirred water layer. Theophylline affects CLi of salicylic acid and quinidine partly by increasing the microclimate pH of the intestine. Theophylline may also affect quinidine CLi by inhibiting the carrier-mediated pathway.     

薯蓣皂苷元丁二酸单酯在大鼠小肠内的吸收动力学

目的 考察薯蓣皂苷元丁二酸单酯(DSAE)在大鼠肠道内的吸收动力学.方法 采用大鼠在体肠吸收方法, 用HPLC测定循环液中的DSAE.结果 DSAE在大鼠小肠内无特定吸收部位,全肠道均有较好吸收.十二指肠、空肠、回肠的每小时吸收百分率(中浓度)分别为13.05%、12.89%、13.13%.不同浓度DSAE在大鼠肠道的吸收速率常数(ka)经方差分析无显著性差异(P＞0.05).结论 DSAE在大鼠小肠的吸收机理可能为被动扩散,在整个肠道均有吸收.     

辛伐他汀大鼠肠吸收动力学研究

目的:研究辛伐他汀在大鼠各肠段的吸收动力学特征。方法:采用大鼠在体肠段回流实验,主要从吸收部位、药物浓度、pH值方面对辛伐他汀的肠段吸收特性进行研究。结果:辛伐他汀在大鼠肠道内无特定吸收部位,各肠段吸收速率常数按十二指肠、结肠、空肠、回肠顺序依次下降,分别为0.03365、0.03190、0.02942、0.02563h-1。辛伐他汀在1.0～20.0μg·mL-1浓度范围内药物吸收量呈线性关系;在pH5.0～7.4内药物吸收不受pH值影响。结论:辛伐他汀在大鼠全肠段均有吸收,吸收符合一级动力学特征,吸收机制为被动扩散,适于制备日服1次缓释给药系统。     

利巴韦林大鼠肠吸收动力学

目的研究利巴韦林在大鼠各肠段的吸收动力学特征。方法应用大鼠在体灌流肠吸收实验考察其吸收动力学 ;采用HPLC法测定利巴韦林在大鼠体内肠吸收回流液中的药物浓度 ;采用UV法测定回流液中酚红浓度的变化。结果利巴韦林在小肠中上部吸收较好 ,吸收速率按十二指肠、空肠、回肠、结肠依次下降 ,吸收速率常数依次为 0 0 970 0、0 0 15 0 0、0 0 0 110、0 0 0 0 0 5h-1;回流液中吸收药物的量与利巴韦林的浓度成正比。结论利巴韦林在肠道的吸收呈一级吸收动力学特征 ,吸收机制为被动吸收 ;主要吸收部位为十二指肠和小肠上部 ,结肠几乎无吸收 ;提示该药适合制成日服 2次的缓释制剂     

Jejunal and ileal absorption of oxprenolol in man: influence of nutrients and digestive secretions on jejunal absorption and systemic availability.

1 Study I evaluated the absorption of oxprenolol in the ileum, compared to jejunum, in healthy volunteers by an intestinal perfusion technique. Around 80 mg of drug were delivered as a saline solution directly in the small bowel. 2 Samples taken 30 cm distally to the site of perfusion showed that 63% of perfused oxprenolol was absorbed in the jejunum and 48% in the ileum; the differences were significant. 3 The plasma concentration-time profiles were similar for the two perfusions. The AUC and Cmax values of free and conjugated oxprenolol for the jejunal perfusion were significantly lower than those of ileum. They showed large but consistent intersubject variations in the two treatments. 4 Study II investigated, using the same technique, the influence of nutrients and digestive secretions on jejunal absorption and systemic availability of this drug. A saline (in treatments A and B) or a nutrient (in treatment C) solution containing oxprenolol was perfused into the jejunum below a balloon either inflated (A) or deflated (B and C). 5 The disappearance rate of oxprenolol from the jejunum was unaffected by endogenous secretions. The mean amount of drug absorbed along a 30-cm jejunal segment accounted for 52 (A) and 57% (B) of the total amount perfused. The intestinal absorption rate was markedly increased in the presence of nutrients (mean amount absorbed 96% for C). 6 The change in the rate of disappearance from the intestine had no effect on the systemic availability of oxprenolol (mean AUC values 8740, 8250 and 8020 nmol l-1 h for A, B and C, respectively) or its elimination from plasma.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of indinavir on the intestinal exsorption of amprenavir,saquinavir and nelfinavir after intravenous administration in rats

To elucidate drug interaction between human immunodeficiency virus (HIV) protease inhibitors (PIs), the effect of indinavir (IDV) on the intestinal exsorption of other HIV PIs, amprenavir (APV), saquinavir (SQV) and nelfinavir (NFV) was investigated in rats using an in situ single perfusion method. IDV in the intestinal perfusate inhibited the exsorption of rhodamine 123 (Rho123), a known P-glycoprotein (P-gp) substrate, from blood into intestinal lumen in a concentration-dependent manner, and the inhibitory potency of 10 micro M IDV in the perfusate was close to that of 10 micro M cyclosporin A (CsA) in the perfusate. Ten micro M of IDV in the intestinal perfusate also decreased significantly the exsorption clearance of Rho123 after intravenous administration. The IDV concentration in this system was not likely to cause hepatic interaction between HIV PIs, because the plasma IDV concentration was far below its inhibition constants for other HIV PIs in the liver microsomes. Thus, 10 micro M of IDV was chosen to investigate the effect of this inhibition on the exsorption of APV, SQV and NFV. IDV in the intestinal perfusate markedly increased the exsorbed amounts of SQV and NFV but not APV after intravenous administrations. Their exsorption clearances, however, showed only a slight increasing tendency or remained unchanged. These findings suggest that in addition to P-gp inhibition, other factors such as CYP3A inhibition might be important in the drug interaction of IDV with APV, SQV and NFV after intravenous administration in rat small intestine. The results obtained in this study will provide useful information to discuss the interactions among PIs when a double protease therapy is used for in HIV-infected patients.     

基于原位单向肠灌流模型研究没食子酸的肠吸收特性

目的考察没食子酸的肠道吸收特性,为提高鞣质类成分生物利用度提供理论依据。方法采用大鼠在体肠单向灌流模型、建立HPLC测定没食子酸的方法,并计算没食子酸在各肠段的吸收速率常数(Ka)及有效表观渗透率系数(Peff);分别研究吸收部位、药物浓度、时间、pH值、P-糖蛋白(P-gp)和多药耐药相关蛋白-2(MRP2)抑制剂对没食子酸吸收的影响。结果没食子酸在不同肠段的Ka顺序为空肠>十二指肠>回肠≈结肠;随着药物浓度的升高,没食子酸的吸收差异无显著性;酸性环境(pH 5.5)有利于没食子酸的吸收;加入P-gp和MRP2抑制剂后,没食子酸的吸收与不加P-gp和MRP2抑制剂比较差异有显著性(P<0.05)。结论没食子酸在大鼠肠道内有较好的吸收,在空肠中吸收最好。初步判断其吸收机制为被动扩散。没食子酸的吸收受P-gp和MRP2的外排影响,可能为P-gp和MRP2底物。     

Drug Exsorption from Blood into the Gastrointestinal Tract

Drugs are exsorbed from the blood across the gastrointestinal membranes by passive or active processes. In the case of a passive transport mechanism, the exsorption of drugs depends on the concentration gradients between the serosal and mucosal sides. The extent of secretion (exsorption) is determined by numerous factors such as extent of binding to serum proteins, distribution volume, lipophilicity, pKa and molecular size of drugs, and the blood flow rate in the gut. Specific transport systems such as P-glycoprotein (P-gp), organic cation and organic anion transporters are found to be involved in active intestinal secretion of drugs. Intestinal secretory transport systems reduce the extent of drug absorption sometimes resulting in low oral bioavailability. It is, therefore, important to know whether poor drug absorption is due to the involvement of specialized secretory transport systems. Modulation of intestinal secretory transport can be a means to enhance absorption of drugs with low oral bioavailability if exsorption of drugs is based on active secretion pathways that are open for control from the "outside.     

Biopharmaceutics classification and intestinal absorption study of apigenin

The aim of the study was to characterize the biopharmaceutics classification system (BCS) category of apigenin (AP) using intrinsic dissolution rate (IDR) and rat intestinal permeability, and to investigate the intestinal absorption mechanism of AP in rats. In the present investigation, equilibrium solubility and intrinsic dissolution rate (IDR) of AP were estimated in phosphate buffers. Effective intestinal permeability (P(eff)) of AP was determined using single-pass intestinal perfusion (SPIP) technique in four segments (duodenum, jejunum, ileum and colon) of rat intestine at three concentrations (10, 50 and 100μg/ml). The aqueous solubility of AP in tested phosphate buffers was very poor with maximum solubility of 2.16μg/ml at pH 7.5. The IDR of AP was very low with a value of 0.006mg/min/cm(2). The minimum and maximum P(eff)s determined by SPIP were 0.198×10(-4) and 0.713×10(-4)cm/s at jejunum and duodenum site, respectively. In addition, the concentration-dependent permeability behavior was observed in the duodenum and jejunum, which suggested that AP was transported by both passive and active carrier-mediated saturable mechanism in these two intestinal segments. However, the observed concentration-independent permeability behavior in ileum and colon indicated primarily passive transport mechanism of absorption of AP in the last two intestinal segments. AP was classified as class II drug of the BCS due to its low solubility and high intestinal permeability. AP could be well absorbed in the whole intestine with the main absorption site at duodenum. The absorption of AP in four intestinal segments exhibited different transport mechanisms.     

盐酸麻黄碱的大鼠肠吸收研究

目的：研究盐酸麻黄碱的肠吸收机制。方法：利用大鼠在体单向肠灌流模型,采用HPLC法测定灌流液中盐酸麻黄碱含量,分别考察灌流速度、盐酸麻黄碱质量浓度、不同肠段以及P-糖蛋白抑制剂对盐酸麻黄碱肠吸收的影响。结果：灌流速度对盐酸麻黄碱吸收速率常数（Ka）和表观吸收系数（Papp）有极显著影响（P〈0.01）;灌流液中盐酸麻黄碱质量浓度对Ka和Papp无显著影响（P〉0.05）;盐酸麻黄碱在小肠各肠段（十二指肠、空肠和回肠）的Ka和Papp无显著性差异（P〉0.05）,但其Ka值显著大于在结肠处的值（P〈0.05）,而各肠段的Papp无显著性差异（P〉0.05）,P-糖蛋白抑制剂对盐酸麻黄碱在各肠段的Ka和Papp无显著影响（P〉0.05）。结论：盐酸麻黄碱在大鼠肠道内的吸收机制为被动扩散,不存在饱和吸收;其在全肠道吸收较好,吸收窗主要在小肠,且小肠内无明显的特定吸收部位;盐酸麻黄碱可能不是P-糖蛋白的底物。     

Modulated pharmacokinetics and increased small intestinal toxicity of methotrexate in bilirubin-treated rats

Objectives The effect of bilirubin treatment on the pharmacokinetics and small intestinal toxicity of methotrexate was evaluated in rats, since bilirubin and its glucuronide conjugates can suppress multidrug resistance‐associated protein‐mediated transport. Methods Rats were treated intravenously with bilirubin and the various clearances and tissue distribution of methotrexate were estimated under a steady‐state plasma concentration. Intestinal toxicity induced by methotrexate was also evaluated by measuring the leakage of alkaline phosphatase (ALP) activity. Probenecid, an inhibitor for multidrug resistance‐associated protein and organic anion transporters, was used for comparison. Key findings The treatment with bilirubin increased the steady‐state plasma concentration and reduced biliary excretion clearance, urinary excretion clearance and intestinal exsorption clearance of methotrexate, as did treatment with probenecid. The intestinal absorption and jejunum distribution of methotrexate also significantly increased in bilirubin‐ and probenecid‐treated rats. A greater leakage of ALP activity to the luminal fluid, with a lower ALP activity in the intestinal mucosal membrane after intestinal perfusion of methotrexate, was observed in bilirubin‐ and probenecid‐treated rats. Conclusions Hyperbilirubinemia, which is involved under various disease states, may increase the small intestinal accumulation and toxicities of methotrexate, since high plasma concentrations of conjugated bilirubin can suppress the function of multidrug resistance‐associated proteins, which facilitate the efflux of methotrexate out of cells.     

酒石酸锑钾口服后胃肠道反应及吸收的规律、影响其吸收的因素

本文选用犬不同部位肠瘘给药法,比较肠管不同部位对酒石酸锑钾(以下简称T.E.)刺激敏感性的差异。用犬肠瘘给药测末梢血锑含量及在位肠襻给药测肠襻中残存锑量法,分析肠管不同部位对T.E.的吸收能力,并观察了大白鼠肠腔经酸化后对T.E.吸收的影响。实验结果:肠管不同部位对T.E.刺激的敏感性及吸收能力均有明显的差别。十二指肠及空肠对T.E.刺激敏感,呕泻反应强烈而频繁,但对T.E.的吸收量较多。迴肠下段及结肠对T.E.刺激感受迟钝,反应轻,次数少,仅有排粪(非泻)无呕吐,对T.E.的吸收量亦比肠管上段显著为低。人工降低肠pH,可显著提高肠管对T.E。的吸收量。     

A new physiologically based, segregated-flow model to explain route-dependent intestinal metabolism.

Processes of intestinal absorption, metabolism, and secretion must be considered simultaneously in viewing oral drug bioavailability. Existing models often fail to predict route-dependent intestinal metabolism, namely, little metabolism occurs after systemic dosing but notable metabolism exists after oral dosing. A physiologically based, Segregated-Flow Model (SFM) was developed to examine the influence of intestinal transport (absorption and exsorption), metabolism, flow, tissue-partitioning characteristics, and elimination in other organs on intestinal clearance, intestinal availability, and systemic bioavailability. For the SFM, blood flow to intestine was effectively segregated for the perfusion of two regions, with 10% reaching an absorptive layer-the enterocytes at the villus tips of the mucosa where metabolic enzymes and the P-glycoprotein reside, and the remaining 90% supplying the rest of the intestine (serosa and submucosa), a nonabsorptive layer. The traditional, physiologically-based model, which regards the intestine as a single, homogeneous compartment with all of the intestinal blood flow perfusing the tissue, was also examined for comparison. The analytical solutions under first order conditions were essentially identical for the SFM and traditional model, differing only in the flow rate to the absorptive/removal region. The presence of other elimination organs did not affect the intestinal clearance and bioavailability estimates, but reduced the percentage of dose metabolized by the intestine. For both models, intestinal availability was inversely related to the intrinsic clearances for intestinal metabolism and exsorption, and was additionally affected by both the rate constant for absorption and that denoting luminal loss when drug was exsorbed. However, the effect of secretion by P-glycoprotein became attenuated with rapid absorption. The difference in flow between models imparted a substantial influence on the intestinal clearance of flow-limited substrates, and the SFM predicted markedly higher extents of intestinal metabolism for oral over i.v. dosing. Thus, the SFM provides a physiological view of the intestine and explains the observation of route-dependent, intestinal metabolism.     

水飞蓟宾在大鼠小肠中的吸收特性

目的考察水飞蓟宾在大鼠各肠段的吸收。方法以一定浓度的水飞蓟宾溶液作为灌流液，以0.1 mL·min^-1进行大鼠不同肠段的单向灌流,于不同时间收集肠灌流液并肝门静脉取血，分别用HPLC测定灌流液和血中药物浓度。结果实验结果表明190 μg·mL^-1水飞蓟宾在不同肠段的吸收速率常数(k_a)和有效透过系数(P_eff)是十二指肠>空肠>回肠>结肠。质量浓度为80 μg·mL^-1的水飞蓟宾灌流液在十二指肠的吸收与190和300 μg·mL^-1的吸收情况均有显著性差异(P<0.05)，但质量浓度为190 μg·mL^-1与300 μg·mL^-1的吸收之间无显著性差异(P>0.05)。肝门静脉血中药物分析也显示十二指肠>空肠>回肠>结肠。结论水飞蓟宾在小肠全肠道均有吸收且有高浓度饱和现象。