5-氟尿嘧啶口服微乳的制备及其大鼠肠吸收作用研究

目的:制备5-氟尿嘧啶(5-Fu)口服微乳,并考察其在大鼠肠吸收的作用。方法:以肉豆蔻酸异丙酯为油相、单辛/癸酸甘油酯为乳化剂,借助伪三元相图法对不同5-Fu微乳处方进行评价;用外翻肠囊法制备肠吸收离体模型,考察5-Fu微乳的吸收部位和促吸收效果。结果:选择肉豆蔻酸异丙酯-单辛/癸酸甘油酯-无水乙醇-水(Km=1∶2)体系作为5-Fu微乳的载药体系;与其溶液比较,5-Fu微乳可明显改善药物的肠吸收,小肠中后段是其最佳吸收部位,90min时累积吸收率微乳是溶液的3倍。结论:所制备的5-Fu微乳性质稳定、肠吸收效果良好。     

Regional Delivery of Model Compounds and 5-Fluorouracil to the Liver by Their Application to the Liver Surface in Rats: Its Implication for Clinical Use

Purpose The purpose of this study was to examine drug distribution in the liver after drug application to the rat liver surface.Methods Phenolsulfonphthalein (PSP) and fluorescein isothiocyanate dextran (MW 4400, FD-4) as model compounds or 5-fluorouracil (5-FU) was applied to the rat liver surface by employing a cylindrical diffusion cell (i.d. 9 mm, 0.64 cm²). Then, blood and the remaining solution in the diffusion cell were collected at selected times, followed by excision of the liver. The excised liver was divided into three sites: the region under the diffusion cell attachment site (site 1), the applied lobe except for site 1 (site 2), and non-applied lobes (site 3).Results In the case of i.v. administration, there were no differences in PSP concentrations among the three sites of the rat liver, and the concentrations rapidly decreased. On the other hand, the PSP concentration in site 1 after application to the rat liver surface was considerably higher than in site 2 and site 3. In addition, the area under the curve (AUC) value (AUC_site1), calculated from the PSP concentration profile in site 1, was about 10 times larger than that in site 3. A similar trend of regional delivery advantage by liver surface application was observed in the case of the macromolecule model FD-4, with a marked AUC_site1 of about 5 times larger than the other two sites. Moreover, we clarified that the anticancer drug 5-FU preferentially distributed in site 1 after application to the rat liver surface.Conclusion These results demonstrate the possibility of regional delivery of drugs to the liver by application to the liver surface.     

Role of Intestinal First-Pass Metabolism of Baicalein in its Absorption Process

Purpose The aim of the present study was to investigate the role of intestinal first-pass metabolism of baicalein (B) in its absorption process.Methods The intestinal absorption of B was characterized using Caco-2 cell monolayer model and rat in situ single-pass intestinal perfusion model. In addition, preliminary metabolic kinetics of B was evaluated in both rat and human intestinal S9 fractions.Results B was well absorbed and extensively metabolized to baicalin (BG), baicalein-7-O-β-glucuronide, in rat intestinal perfusion model, whereas less extent of metabolism was observed in the Caco-2 cell monolayer model. Moreover, BG generated in the intestinal epithelium during the absorption of B also rapidly transported to both the apical side (the apical chamber of Caco-2 model and the perfusate of the intestinal perfusion model) as well as the basolateral side of the small intestine (the basal chamber of Caco-2 model and the mesenteric vein of the intestinal perfusion model). From the preliminary metabolic studies, it was found that a higher loading dose of B resulted in a less extent of metabolism in intestine. In addition, the extent of metabolism of B was similar in jejunum and ileum when 50 μM of B was perfused through different sections of rat small intestine.Conclusion The first-pass metabolism of B in small intestine may play an important role in its low oral bioavailability.     

Simultaneous Prediction of Intestinal Absorption and Metabolism Using the Mini-Ussing Chamber System

The purpose of this study was to investigate the possibility of simultaneous prediction of the intestinal absorption and metabolism in a mini-Ussing chamber equipped with rat intestinal tissues, based on the transport index (TI). TI value was defined as the sum of drug amounts, by mass balance method, transported to the basal-side component and drug amounts accumulated in the tissue, which are normalized by area under the curve of the drug in the apical compartment. Midazolam and nifedipine with high permeability were used as typical P450 substrates to examine the possibility of simultaneous prediction of intestinal absorption and metabolism. The metabolite formation of both compounds was observed and ketoconazole strongly inhibited the metabolite formation of both compounds in rat intestinal tissues, leading to the improvement of the TI value to a statistically significant extent for both compounds. TI ratio of nifedipine between in the presence and absence of ketoconazole was larger than that of midazolam, which was consistent with the reported lower value of fraction absorbed multiplied by intestinal availability of nifedipine. Therefore, the mini-Ussing chamber, equipped with animal intestinal tissues, showed potential to predict the intestinal absorption and metabolism simultaneously.     

Intestinal Absorption of Miltefosine: Contribution of Passive Paracellular Transport

Purpose This study aimed to characterize the transepithelial transport of miltefosine (HePC), the first orally effective drug against visceral leishmaniasis, across the intestinal barrier to further understand its oral absorption mechanism. Materials and Methods Caco-2 cell monolayers were used as an in vitro model of the human intestinal barrier. The roles of active and passive mechanisms in HePC intestinal transport were investigated and the relative contributions of the transcellular and paracellular routes were estimated. Results HePC transport was observed to be pH-independent, partially temperature-dependent, linear as a function of time and non-saturable as a function of concentration. The magnitude of HePC transport was quite similar to that of the paracellular marker mannitol, and EDTA treatment led to an increase in HePC transport. Furthermore, HePC transport was found to be similar in the apical-to-basolateral and basolateral-to-apical directions, strongly suggesting that HePC exhibits non-polarized transport and that no MDR-mediated efflux was involved. Conclusions These results demonstrate that HePC crosses the intestinal epithelium by a non-specific passive pathway and provide evidence supporting a concentration-dependent paracellular transport mechanism, although some transcellular diffusion cannot be ruled out. Considering that HePC opens epithelial tight junctions, this study shows that HePC may promote its own permeation across the intestinal barrier.     

Electrorepulsion Versus Electroosmosis: Effect of pH on the lontophoretic Flux of 5-Fluorouracil

Purpose. To delineate the contributions of electrorepulsion and electroosmosis to the iontophoretic flux of 5-FU across porcine skin in vitro. Also, the isoelectric point (pI) of the skin model was determined. Methods. The electrotransport of 5-FU, anode-to-cathode ('anodal') and cathode-to-anode ('cathodal') was determined as a function of the pH of the electrolyte bathing the skin. Results. At pH 8.5, the drug (pK_a 8) is negatively charged and 'cathodal', viz. electrorepulsive, transport is much greater than that in the opposite direction. At pH 7.4, where 25% of 5-FU is charged, electrorepulsive and electroosmotic ('anodal') fluxes are balanced. Decreasing the pH to 6, and then 5, reduces the percentage of ionized 5-FU such that 'anodal' electroosmosis dominates across the negatively-charged membrane. But, at pH 4, 'anodal' and 'cathodal' fluxes are again equal suggesting neutralization of the skin (i.e., pI 4). This is confirmed at pH 3, where 'cathodal' electroosmosis dominates across the now net-positively charged barrier. Conclusions. Electrotransport is sensitive, mechanistically, to the properties of the permeant and of the skin; interactions of, for example, the drug or constituents of a formulation, that alter the barrier's net charge, can affect iontophoretic delivery. The pI of porcine ear skin is 4.     

Regulation of Paracellular Absorption of Cimetidine and 5-Aminosalicylate in Rat Intestine

Purpose. Isolating the relative contributions of parallel transcellular and paracellular transport to the intestinal absorption of small hydrophilic molecules has proven experimentally challenging. In this report, lumenal appearance of drug metabolite is utilized as a tool to assess the contribution of paracellular transport to the absorption of cimetidine and 5-aminosalicylate (5ASA) in rat small intestine. Methods. Steady-state intestinal absorption and elimination of cimetidine and 5ASA were studied in single-pass intestinal perfusions in rats. Results. Both drugs were metabolized in intestinal epithelia with subsequent metabolite secretion into the intestinal lumen. Jejunal cimetidine absorption decreased with increasing perfusion concentration while the ratio of lumenal metabolite to lumenal drug loss increased. Cimetidine uptake at perfusion concentrations above 0.4 mM resulted in over 80% drug elimination into the jejunal lumen. Inhibition of intracellular metabolism of cimetidine by methimazole did not alter epithelial uptake but totally abolished transepithelial cimetidine flux indicating an elevation of intracellular cimetidine. Similarly, co-perfusion of 5ASA with cimetidine and methimazole totally abolished 5ASA absorption but increased lumenal levels of N-acetyl 5ASA indicating an increase in intracellular uptake of 5ASA. Conclusions. Cimetidine and 5ASA absorption across rat jejunal epithelia are exclusively paracellular. Elevation of intracellular cimetidine, inferred from mass balance considerations, restricts paracellular transport of both drugs.     

The Enhancer Effect of Several Phenyl Alcohols on Percutaneous Penetration of 5-Fluorouracil

Pharmaceutical Research -     

Mechanism and Site Dependency of Intestinal Mucosal Transport and Metabolism of Thymidine Analogues

This study has been undertaken to investigate the mechanisms of intestinal mucosal transport and metabolism of thymidine analogues and to identify any optimal site(s) of the rat intestine particularly involved in the absorption of thymidine analogues. The intestinal absorption of 3-azido-3-deoxythymidine (AZT) was studied at three initial concentrations in four segments of the rat intestine using an in situ recirculating perfusion technique. Disappearance of AZT followed first-order kinetics throughout the gastrointestinal (GI) tract at all tested concentrations. The apparent first-order rate constants were found to be relatively invariant over a broad range of concentrations from 0.01 to 1.0 mM. Corrected for the length of each segment, the apparent permeability (P _app) of AZT was 3.01 ± 0.32 × 10^–5 cm/sec (mean ± SE) in the duodenum, 2.06 ± 0.24 × 10^–5 cm/sec in the upper jejunum, 0.76 0.13 × 10^–5 cm/sec in the combined lower jejunum and ileum, and 0.32 ± 0.10 × 10^–5 cm/sec in the colon, which indicated that intrinsic absorptivity was greater in the upper GI tract than in the lower portions possibly due to the differences in surface area for absorption. No AZT metabolite appeared in any part of the GI tract. On the other hand, thymidine and other analogues, i.e., 5-iodo-2-deoxyuridine and 2-deoxyuridine, were rapidly metabolized into nucleobase and sugar in the upper GI tract, whereas in the colon no metabolite appeared. A free 3-OH group appears to be necessary for the metabolism (catabolism) of thymidine analogues in the rat intestine mainly by pyrimidine nu-cleoside phosphorylase. Finally, bile salt-acylcarnitine mixed micelles appeared to be an effective adjuvant in promoting colonic absorptions of AZT and phenol red. The use of mixed micelles increased the apparent permeabilities of AZT in the colon by a factor of 5.4, and for phenol red the permeability increased from a negligible value to 1.76 × 10^–5 cm/sec. Since the absorptions of both AZT and phenol red were enhanced by mixed micelles, a paracel-lular transport pathway may be involved.     

Intestinal Peptide Transport Systems and Oral Drug Availability

The intestinal peptide transport system has broad substrate specificities. In addition to its physiological function of absorbing di- and tripeptides resulting from the digestion of dietary proteins, this transport system also absorbs some orally administered peptidomimetic drugs, including -lactam antibiotics, angiotensin converting enzyme inhibitors, renin inhibitors, bestatin, thrombin inhibitors, and thyrotropin-releasing hormone and its analogues. There have been several studies on the mechanism and substrate structure-affinity relationship for this transport system. Rapid progress has been made recently in studies on the molecular basis of the intestinal peptide transport system. A protein apparently involved in peptide transport has been isolated from rabbit small intestines, and genes for human intestinal peptide transporters have been cloned, sequenced and functionally expressed. This review summarizes these studies and addresses the pharmaceutical potential of the intestinal peptide transport system.     

Determination of Transport Rates for Arginine and Acetaminophen in Rabbit Intestinal Tissues in Vitro

The in vitro Ussing technique was employed to examine transport rates for acetaminophen and arginine across rabbit intestinal tissues. Mannitol and transepithelial conductance were used to monitor the integrity of rabbit intestinal tissues and the basal and stimulated short-circuit current were measured to assess functional viability. Transepithelial transport of acetaminophen, arginine, and mannitol was determined in rabbit jejunum, ileum, and distal colon. Transepithelial transport of arginine in the ileum and jejunum was composed of both passive (nonsaturable) (P _m = 0.06) and saturable components (K _T = 0.6-0.7 mM; J _max = 0.3-0.4 µmol/hr · cm²). The saturable component of arginine fluxes was abolished by pretreatment of the tissue with serosal ouabain (0.1 mM). In the distal colon, both unidirectional arginine fluxes were nonsaturable. In the segments examined, both unidirectional fluxes of acetaminophen were nonsaturable over the concentration range from 0.1 to 30 mM. These results provide values for maximal permeabilities attained by molecules traversing both the cellular and the paracellular pathways and, by comparison to their in vivo bioavailabilities, provide selection criteria for evaluating drug candidates for oral activity.     

Intestinal Drug Absorption and Metabolism in Cell Cultures: Caco-2 and Beyond

Pharmaceutical Research -     

包裹5-氟尿嘧啶的二氧化硅纳米颗粒的制备及细胞毒性研究

目的 制备包裹5-氟尿嘧啶的二氧化硅(5-Fu/SiO₂)纳米颗粒,并对其药剂学参数及体外细胞毒性进行研究。方法 利用反相微乳化法制备5-Fu/SiO₂纳米颗粒,对5-Fu投入量及反应时间等合成条件进行优化,考察纳米颗粒的稳定性及体外释药行为,并采用MTT法对其体外细胞毒性进行研究。结果 当5-Fu投入量为1.33 mg及反应时间为24 h时,5-Fu/SiO₂纳米的载药率及包封率达到最高,分别为1.03%及24.77%,同时该纳米在48 h内释放率达到41.31%,7 d内粒径无明显变化。细胞毒性实验表明,5-Fu/SiO₂纳米对人肝癌细胞具有明显的抑制效果,而空白SiO2纳米对细胞活性影响较小。结论 成功制备了稳定性好、缓释时间长的5-Fu/SiO₂纳米颗粒,为开发5-Fu缓释剂型提供了新的思路。     

HPLC法快速测定5—氟尿嘧啶血药浓度

目的 :建立反相高效液相色谱法测定人血清中 5 -氟尿嘧啶浓度。方法 :血清样品用乙酸乙酯提取 ,水浴氮气吹干 ,残留物用水溶解后进样。色谱柱为C1 8柱 (2 5 0mm× 4 6mm) ,水为流动相 ,流速 1 0mL·min- 1 ,紫外检测波长 2 73nm ,桂皮酸为内标。结果 :本法最低检测浓度为 0 0 5 μg·mL- 1 ,线性范围为 1 0～ 5 0 0 μg·mL- 1 ,日内RSD为 2 7%～ 4 1% ,日间RSD为 3 8%～ 4 7(n =4)。结论 :该法适用于 5 -氟尿嘧啶的药代动力学研究及临床血药浓度检测。试验结果表明 ,该方法经济、简便、快速、灵敏度高、重现性好。     

5-氟尿嘧啶-蛇葡萄素复方脂质体的制备

目的制备5-氟尿嘧啶-蛇葡萄素复方脂质体并建立同时测定2种药物含量的紫外分光光度法。方法紫外分光光度法确定5-氟尿嘧啶和蛇葡萄素的测定条件。比较几种脂质体的制备方法,以包封率和粒径为指标,确定最优制备方法;采用正交设计进行优化处方和制备工艺。结果采用紫外分光光度法测定5-氟尿嘧啶和蛇葡萄素,2种药物的加样回收率均在99%~102%之间。以薄膜分散超声法为制备方法,以单因素考察结合正交设计优选出的最佳处方和制备工艺,主药∶卵磷脂=1∶20,卵磷脂∶胆固醇=4∶1,磷脂浓度50 g.L 1;维生素E的用量为5%,磷酸盐缓冲溶液pH为7.4。5-氟尿嘧啶和蛇葡萄素的包封率分别为(44.79±1.55)%和(75.47±0.91)%(n=3);复方脂质体的粒径为(142±3.6)nm。5-氟尿嘧啶和蛇葡萄素的体外12 h累积释放率分别为43.05%和60.24%。结论将5-氟尿嘧啶和蛇葡萄素同时包封于脂质体制备成复方脂质体,所采用的制备工艺简单可行,重复性好,包封率较高。建立的紫外分光光度法可同时测定复方脂质体中的5-氟尿嘧啶和蛇葡萄素含量。     

Mechanisms of 5-Fluorouracil (5-FU) Transport in Isolated Rabbit Choroid Plexus Tissue Slices

Pharmaceutical Research -     

Intestinal First-Pass Metabolism of Eperisone in the Rat

Purpose. The purpose of this study was to clarify quantitatively the contribution of the intestine to the first-pass metabolism of eperisone in rats. Methods. The systemic availabilities of eperisone were estimated by administering the drug into the duodenum, portal vein, and femoral vein in rats in vivo. The first-pass metabolism of eperisone was confirmed in the perfused rat small intestine in situ. Metabolism of eperisone to an -1-hydroxylated metabolite (HMO), the first step of eperisone metabolism, was studied using rat intestinal microsomes in vitro. Results. The bioavailabilities in the intestine were 0.176 and 0.0879 at administration rates of 100 and 25 mg/h/kg, respectively, whereas those in the liver were 0.532 and 0.486, respectively. In the intestinal perfusion experiment, the appearance clearance to the portal vein from the intestinal lumen was much lower than the elimination clearance from the intestinal lumen, resulting in high metabolic clearance of eperisone in the small intestine. Eperisone was biotransformed to HMO by rat intestinal microsomes, and this was inhibited by -naphthoflavone and an anti-rat CYP1A antibody. Conclusions. Those data strongly suggest that eperisone may be metabolized to HMO by CYP1A in rat intestinal microsomes during the first-pass through the epithelium of the small intestine.     

5-氟尿嘧啶乙醇脂质体的改性及其透皮吸收研究

目的:研究胆固醇对5-氟尿嘧啶(5-FU)乙醇脂质体的改性及其体外透皮扩散的影响。方法:制备不同胆固醇含量的5-FU乙醇脂质体,并考察胆固醇含量对脂质体粒径、Zeta电位、分散指数、包封率、皮内药物滞留量等指标的影响。结果:加入胆固醇后粒径和Zeta电位变化不大,分散指数从0.584降至0.143,5-FU包封率从28.6%增至48.8%,皮内残留5-FU量从40%增至80%以上。结论:胆固醇不会改变乙醇脂质体的粒径大小及Zeta电位,但可提高其分散性和稳定性;加入适量的胆固醇可提高乙醇脂质体中5-FU的包封率及皮内药物滞留量。     

羧基化碳纳米管 醋氯芬酸的离体肠吸收特性变化研究

目的：研究羧基化多壁碳纳米管负载醋氯芬酸（AC）的制备工艺,以及负载物（AC-MWCNTs-COOH）的肠吸收特性.方法：采用冷冻球磨法制备负载物,采用FIR、紫外光谱进行表征;采用离体外翻肠囊模型研究AC及负载物在不同肠段的吸收特性,采用HPLC法测定样品浓度,计算吸收速率常数（Ka）.结果：AC原料与负载物均随着药液中AC浓度上升,Ka呈线性增加;负载物的Ka均低于AC原料.AC原料在各个肠段的Ka差异均无统计学意义（P＞0.05）,而负载物在十二指肠的Ka大于空肠、回肠、结肠,后三者差异无统计学意义（P＞0.05）;负载物浓度为0.54 mg· ml-1时,各肠段的Ka依次为0.740 6、0.129 2、0.181 3、0.241 1μg·h-1·cm-2.结论：负载物具有缓释效果,在小肠内的Ka与原料相比发生改变;在小肠中均为被动扩散吸收.