乳化蒸发-低温固化法制备槲皮素纳米脂质体及其在大鼠血浆存在形式与脑靶向性

目的:制备槲皮素纳米脂质体(QUE-NL),考察槲皮素在大鼠血浆中存在的物质形式及其脑内分布特性。方法:采用乳化蒸发-低温固化法制备槲皮素纳米脂质体;采用高效液相色谱法检测大鼠血浆中槲皮素存在形式及脑组织中槲皮素浓度。结果:槲皮素在大鼠血浆中主要是以槲皮素苷元和异鼠李素存在,且槲皮素纳米脂质体在大鼠脑内吸收分布优于槲皮素原料药和普通脂质体。结论:吸收进入血浆中的槲皮素主要是以结合形式而非单体形式存在,以乳化蒸发-低温固化法制备的槲皮素纳米脂质体,能显著促进槲皮素在大鼠脑内的吸收,可为进一步应用于脑胶质瘤的治疗提供实验依据。     

槲皮素长循环纳米脂质体的小鼠口服吸收研究

目的 制备槲皮素长循环纳米脂质体(QUE-LCL),并考察其理化性质及小鼠口服吸收特性. 方法 以山嵛酸甘油酯和聚山梨酯-80为载体材料,采用乳化蒸发-低温固化法,以正交实验法优化的处方与制备工艺制备槲皮素长循环纳米脂质体;透射电镜观察其微观形态,并测定其粒径分布;建立高效液相色谱法测定纳米脂质体和胃肠道内容物及粪样混合物中槲皮素含量. 结果 槲皮素长循环纳米脂质体为球状或类球状,平均粒径为172.63 nm,包封率为91.77%,其小鼠体内吸收优于槲皮素原料药和槲皮素普通脂质体. 结论 乳化蒸发 低温固化法适于制备槲皮素长循环纳米脂质体,以正交优化法制备的制剂促进小鼠对槲皮素的口服吸收.     

双氯芬酸钠柔性纳米脂质体的制备及离体透皮扩散的研究

目的：制备双氯芬酸钠柔性纳米脂质体并研究柔性纳米脂质体对双氯芬酸钠离体透皮扩散速率的影响。方法：用旋转蒸发超声法制备柔性脂质体，采用改良的Franz扩散池进行体外透皮吸收试验，结果：五种处方的柔性纳米脂质体可不同程度地增加双氯芬酸钠的透皮速率，其中处方IV的透皮速率最大，为双氯芬酸钠饱和溶液的8倍，结论：柔性纳米脂质体可显地促进双氯芬酸钠的透皮吸收。     

甲氨蝶呤柔性纳米脂质体凝胶的构建及体外经皮渗透性研究

目的 制备甲氨蝶呤(methotrexate,MTX)柔性纳米脂质体凝胶,并研究其体外经皮渗透行为.方法 采用逆向蒸发法制备MTX柔性纳米脂质体,以卡波姆940为基质制成脂质体凝胶,并考察其初步稳定性;Franz扩散池研究MTX柔性纳米脂质体凝胶与普通凝胶的经皮渗透规律.结果 脂质体凝胶4℃下稳定性良好;体外透皮试验表明,MTX柔性纳米脂质体凝胶的累积透过量明显＜MTX普通凝胶(P＜0.05),皮肤滞留量＞MTX普通凝胶(P＜0.05).结论 MTX柔性纳米脂质体凝胶可显著提高药物的皮肤滞留量,而不增加药物进入血液循环的量,能有效降低药物潜在的全身毒性,有望成为MTX局部治疗的新剂型.     

正交试验优选槲皮素纳米脂质体冻干粉制备工艺

目的制备含有不同冻干保护剂的槲皮素纳米脂质体(QUE-NL)冻干粉,筛选出最佳制备工艺。方法采用正交试验法,以纳米脂质体包封率和载药量等作为评价指标,采用多指标综合分析方法优选槲皮素纳米脂质体冻干粉冻干保护剂的最佳比例。结果槲皮素纳米脂质体冻干粉的最佳制备工艺为水∶油相比例为1.5∶1.0,冻干保护剂选用麦芽糖和甘露醇,5%甘露醇+5%麦芽糖(1∶1),以此制备工艺制得的纳米脂质体冻干粉包封率和载药量较为理想。结论以优选工艺制备槲皮素纳米脂质体冻干粉的方法简易、可行。     

槲皮素纳米脂质体冻干粉针的制备及其质量评价

目的:研究槲皮素纳米脂质体冻干粉针的制备方法,并对其进行初步的质量评价。方法:以乳化蒸发-低温固化法和冷冻干燥法制备含有不同冻干保护剂的槲皮素纳米脂质体(QUE-NL)冻干粉,以包封率为评价指标,对制备工艺和处方进行单因素考察,并考察其理化性质,筛选出最佳配方。并对冻于粉针进行稳定性影响因素试验。结果:该法制得的脂质体包封率较佳;制备过程中,槲皮素纳米脂质体的包封率受药脂比影响较大,受胆固醇磷脂比影响较小;采用5%甘露醇+5%麦芽糖作为冻干保护剂冻干效果更好;所得冻于粉针对温度、光照较为敏感,也易受湿度影响。结论:5%甘露醇+5%麦芽糖是槲皮素纳米脂质体较合适的冻干保护剂,初步的稳定性考察结果表明,槲皮素纳米脂质体冻干粉针宜低温、避光、密封保存。以本试验方法制备的槲皮素纳米脂质体冻干粉粒径较小,包封率高,稳定性好,制备工艺合理可行。     

槲皮素长循环纳米脂质体的制备与质量评价

目的 制备槲皮素长循环纳米脂质体,并进行质量评价.方法 采用乳化蒸发-低温固化法制备槲皮素长循环纳米脂质体;以形态、粒径、包封率和载药量等作为纳米脂质体质量评价指标.结果 采用乳化蒸发-低温固化法制备的槲皮素长循环纳米脂质体平均粒径为172.63 nm,包封率为85.90%,载药量为23.55%.结论 该试验制备的槲皮...     

低分子肝素微乳及其纳米脂质体作大鼠口服抗凝效果的比较

目的　制备低分子肝素纳米脂质体及微乳制剂,并进行大鼠口服吸收后抗凝效果的比较。方法　用逆相蒸发法制备低分子肝素纳米脂质体;按三元相图摸索微乳区;以涡旋振荡制备低分子肝素微乳,马尔文激光粒度分析仪测量粒径,以大鼠用药前后血液凝固时间的变化研究其口服吸收的效果。结果　低分子肝素纳米脂质体平均粒径为84 . 1nm ,包封率为2 3 .6 %,微乳平均粒径5 2 . 8nm ;大鼠灌胃微乳及脂质体后,血液凝固时间显著延长。结论　纳米脂质体及微乳对低分子肝素大鼠口服吸收均有良好的促进作用,微乳效果好于纳米脂质体。     

紫杉醇纳米脂质体凝胶剂的制备及体外透皮研究

目的制备紫杉醇纳米脂质体凝胶剂,考察其粒径、粒径分布、包封率、体外释放度及透皮特性。方法采用薄膜蒸发高压微射流法制备紫杉醇纳米脂质体,以卡波姆为凝胶基质,研制紫杉醇纳米脂质体凝胶剂,采用正交试验探索最佳工艺。用粒径测定仪测定脂质体的粒径及其粒径分布,低速-超速相结合法测定包封率,透析膜扩散法进行体外释放试验,以离体小鼠皮结合改良Franz扩散装置考察其体外透皮特性。结果紫杉醇纳米脂质体的最佳工艺:卵磷脂的含量为2%,药物与磷脂质量比为1∶30,磷脂与胆固醇的质量比为10∶1。测得的粒径为81.8 nm;粒径分布系数为0.180;平均包封率73.2%。纳米脂质体凝胶剂72 h累积释放百分率为79.04%;48 h的单位面积累积渗透量为429.68μg·cm?2。结论该制剂制备工艺简单,易于涂布,具有较高的包封率,粒径较小且分布均匀,体外释放缓慢。纳米脂质体能促进脂溶性药物紫杉醇透过皮肤。     

环孢素脂质微粒经小鼠皮肤给药的渗透机理研究

目的　研究普通纳米脂质体、柔性纳米脂质体及胆酸钠-磷脂混合胶团对环孢素经小鼠皮肤给药的渗透机理。方法　将含药载体非封闭性应用于离体或在体小鼠皮肤,测定了皮肤、接收介质和血液中环孢素含量。结果 离体条件下,应用柔性纳米脂质体和混和胶团后均能在接收液中检测到药物,但普通脂质体却不能。在体条件下,应用柔性纳米脂质体后8 h血药浓度到达峰值,而应用普通脂质体和混合胶团后在血中几乎未检测到药物。结论 柔性纳米脂质体在皮肤水合压力下发生变形,携药透过皮肤;普通脂质体主要与皮肤发生融合产生蓄积作用;混合胶团在水溶液状态下发挥其渗透促进作用。     

金丝桃苷在Caco-2细胞模型中的跨膜转运方式

目的研究金丝桃苷在Caco-2细胞模型中的吸收机制。方法用Caco-2细胞单层模型研究金丝桃苷的双向转运,考察pn、药物质量浓度、方向、温度、抑制剂对金丝桃苷细胞转运的影响。采用HPLC法检测金丝桃苷的含量,计算其表观渗透系数(P_app)。结果金丝桃苷的细胞转运P_app。具有pH依赖性。金丝桃苷肠腔（A）侧→基底（B）侧P_app>B→A,并且随着金丝桃苷质量浓度的增大而减小,具有浓度依赖性。P-gp抑制剂维拉帕米增加金丝桃苷的细胞正向转运P_app,降低了其逆向转运P_app。金丝桃苷较高浓度时,MRPl抑制剂吲哚美辛和ATP抑制剂叠氮化钠显著降低了金丝桃苷的转运量。结论金丝桃苷在Caco-2细胞单层模型中的转运具有pH依赖性和浓度依赖性,是以主动转运为主,被动扩散为辅,同时涉及外排蛋白作用的转运方式。     

蜕皮甾酮在Caco-2细胞模型中的摄取和跨膜转运研究

目的以Caco-2细胞单层模型,首次研究了蜕皮甾酮的口服吸收与转运特性。方法采用普通Caco-2细胞模型、表达活性CYP3A4酶的Caco-2细胞模型,分别从AP→BL方向和BL→AP方向研究蜕皮甾酮的摄取和跨膜转运规律。结果蜕皮甾酮在2种模型中的表观渗透系数(Papp)在0.1×10-6～1×10-6cm.s-1之间,药物吸收情况为1%～10%;在4 h的实验过程中,4种浓度蜕皮甾酮的ER值均小于1.5。结论研究表明,蜕皮甾酮主要以被动扩散的方式被细胞摄取和转运,其跨膜转运特性未受到CYP3A4-介导机制的影响。     

Studies on the transport of thyrotropin-releasing hormone (TRH) analogues in Caco-2 cell monolayers

The transport mechanisms of thyrotropin-releasing hormone (TRH) and its pharmacologically active analogues ((3-methyl-His(2))TRH (MeTRH), taltirelin, montirelin, azetirelin) across Caco-2 cell monolayers were characterized. The results of kinetic analysis showed a linear relationship between the concentration (over the range 0.5-10 mM) and apical-to-basolateral transport rate of these agents. The permeability coefficients (P(app)) of these agents were not substantially different from each other, and their P(app) ratios of the basolateral-to-apical over the apical-to-basolateral transport were close to one (0.73-1.23). The cellular transport of [(3)H]MeTRH at low concentrations (3-15 nM) showed a linear relationship between the concentration and transport rate. The transport of [(3)H]MeTRH in Caco-2 cell monolayers was neither affected by TRH nor TRH analogues, and there was little difference in P(app) values between [(3)H]MeTRH and [(14)C]mannitol. The cell-per-medium ratio of [(3)H]MeTRH in the cellular uptake experiment was similar to the value of [14C]mannitol. A large excess of TRH and MeTRH did not significantly influence cell-per-medium ratios of [(3)H]MeTRH in Caco-2 cell monolayers. The k'(IAM) value, which represents lipophilicity, was decreased in the following order: montirelin > taltirelin > TRH > azetirelin, and the values varied from 0.234 to 1.028. These results indicate that a paracellular passive diffusion may be the major route for the transport of TRH and its analogues in Caco-2 cell monolayers.     

Permeability of 5-fluorouracil and its prodrugs in Caco-2 cell monolayers: evidence for shift from paracellular to transcellular transport by prodrug formation

The membrane transport properties of 5-FU and its prodrugs were investigated using Caco-2 cell monolayers. To elucidate a membrane transport of prodrug itself, 2-O-alkoxy-5-fluoropyrimidone (2-O-alkoxy-5FP) derivative, which was stable in buffer and Caco-2 cell, was selected as a test compound. The permeation of 2-O-alkoxy-5FP derivatives was markedly greater than that of 5-FU. The dose-dependency of apical to basal (AP-BL) and BL-AP transports showed that 5-FU was dominantly transported by passive diffusion and ethoxy-5FP was transported by passive diffusion and unsaturated efflux transporter in the apical membrane of Caco-2 cell monolayers. Furthermore, treatment of Cap-Na, an enhancer of paracellular transport, demonstrated that 5-FU and 2-O-alkoxy-5FP derivatives permeated through paracellular and transcellular route, respectively. The significantly increased permeability of 2-O-alkoxy-5FP derivatives was mainly related to capping of hydrophilic functional group, and the moderate increase in permeability among 2-O-alkoxy-5FP derivatives was dependent on prolongation of the alkyl chain length.     

HT29-MTX and Caco-2/TC7 monolayers as predictive models for human intestinal absorption: role of the mucus layer.

The permeability of 19 compounds in both the Caco-2/TC7 and HT29-MTX models was determined, and the ability of each model to predict intestinal absorption in humans was compared. Similar apparent permeability values (log P(app)) were obtained in both models for the majority of compounds tested, and plots of log P(app) versus fraction absorbed in humans gave comparable sigmoidal curves. A linear correlation was also observed between the log P(app) values derived from these two models, which suggests that HT29-MTX is an alternative model for absorption prediction in humans. The similarity of both the diffusion coefficients and permeability values obtained for a range of hydrophilic and lipophilic compounds in the two models indicates that the mucus layer secreted by the human adenocarcinoma HT29-MTX goblet cells does not constitute a diffusion barrier to such compounds. The lack of P-glycoprotein (P-gp) in the HT29-MTX cell line may explain the higher permeability values obtained for cimetidine and sumatriptan in this model compared with those derived from the Caco-2/TC7 monolayers. The results suggest that the HT29-MTX model can be used to rank order the passive permeability of compounds, irrespective of their potential interaction with P-gp, which may facilitate optimization of the physicochemical features of compounds within a chemical series.     

矢车菊黄素在Caco-2细胞模型中的吸收机制研究

目的 研究矢车菊黄素（centaureidin）在Caco-2细胞单层模型中的吸收机制。方法 以HPLC分析矢车菊黄素浓度,用Caco-2 细胞单层模型评价吸收时间、药物浓度、介质pH值、抑制剂等对矢车菊黄素吸收的影响,研究矢车菊黄素的吸收机制,计算表观渗透系数（apparent permeability coefficient, Papp）。结果 药物的吸收与药物浓度和吸收时间正相关;弱酸性介质条件下有利于药物的吸收;2,4-二硝基酚（DNP）对药物吸收无影响,但异博定（verapamil）可增加药物的吸收;从肠腔侧到基底侧的转运小于基底侧到肠腔侧的转运。结论 矢车菊黄素在Caco-2细胞模型中的吸收主要是被动转运,受P-糖蛋白的外排作用。     

Evaluation of transport mechanism of prodrugs and parent drugs formed by intracellular metabolism in Caco-2 cells with modified carboxylesterase activity: temocapril as a model case

The intestinal absorption mechanism of temocapril, an ester-type prodrug of temocaprilat, was evaluated using Caco-2 cell monolayers with or without active carboxylesterase (CES)-mediated hydrolysis. The inhibition of CES-mediated hydrolysis was achieved by pretreatment of the monolayers with bis-p-nitrophenyl phosphate (BNPP), which inhibited 94% of the total hydrolysis of temocapril in the Caco-2 cells. The remaining 6% hydrolysis was due to the presence of serine esterases, other than CES, on the cell membranes. Transport experiments under CES-inhibited conditions showed temocapril not to be a substrate for peptide transporter 1 (PEPT1) or organic anion transporting polypeptides (OATPs), but to be an inhibitor of PEPT1; P-glycoprotein (P-gp) and breast-cancer-resistant protein (BCRP) were responsible for the efflux of temocapril, which was mainly absorbed by passive diffusion at low apical pH. In Caco-2 cell monolayers with CES-mediated hydrolysis intact, temocaprilat derived from temocapril, was 2.5-fold more rapidly transported into the apical compartment than into the basolateral compartment due to the presence of microvilli on the apical membrane. In contrast, temocaprilat at low intracellular concentrations, was preferentially transported across the basolateral membrane under CES-inhibited conditions.     

Caco-2 monolayers in experimental and theoretical predictions of drug transport

This review examines the use of Caco-2 monolayers in the prediction of intestinal drug absorption. First, the different routes of drug transport in Caco-2 monolayers are compared with those seen in vivo. Second, the prediction of drug absorption in vivo from transport experiments in cell monolayers is discussed for different classes of drugs. Finally, the use of Caco-2 monolayers as a reference model in physico-chemical and theoretical predictions of drug absorption is discussed. We conclude that Caco-2 monolayers can be used to identify drugs with potential absorption problems, and possibly also to select drugs with optimal passive absorption characteristics from series of pharmacologically active molecules generated in drug discovery programs.     

High glucose concentration in isotonic media alters Caco-2 cell permeability

Caco-2 cell permeability was evaluated in isotonic media containing high (25mM) or physiological (5.5mM) glucose concentrations. Transepithelial electrical resistance (TEER) and membrane fluidity were measured to assess glucose-induced alterations in physical barrier properties. In parallel, distribution of the actin filament (F-actin) and zonula occludens-1 (ZO-1) proteins was assessed by confocal microscopy. Transepithelial fluxes of mannitol, hydrocortisone, digoxin, and glycyl sarcosine (Gly-Sar) that permeate the intestinal mucosa by various pathways were measured to quantify the effect of glucose-induced changes on Caco-2 cell permeability. High glucose decreased maximum TEER of cell monolayers by 47%, whereas membrane fluidity at the hydrophobic core and lipid/polar head interphase was significantly increased. F-actin distribution in high glucose cells appeared more diffuse while ZO-1 was unchanged. Mannitol and hydrocortisone fluxes across Caco-2 cells cultured in high glucose increased by 65% and 24%, respectively. In addition, high glucose decreased the maximum transport capacity (Vmax) of PepT-1. P-glycoprotein activity, however, was unchanged. In conclusion, high extracellular glucose concentration in isotonic media significantly alters physical barrier properties of Caco-2 cell monolayers, which predominantly affects transepithelial transport of solutes permeating the cell barrier by paracellular and transcellular passive diffusion and facilitated transport mediated by the proton-dependent oligopeptide transporter (PepT-1).     

Caco-2 monolayers in experimental and theoretical predictions of drug transport

This review examines the use of Caco-2 monolayers in the prediction of intestinal drug absorption. First, the different routes of drug transport in Caco-2 monolayers are compared with those seen in vivo. Second, the prediction of drug absorption in vivo from transport experiments in cell monolayers is discussed for different classes of drugs. Finally, the use of Caco-2 monolayers as a reference model in physico-chemical and theoretical predictions of drug absorption is discussed. We conclude that Caco-2 monolayers can be used to identify drugs with potential absorption problems, and possibly also to select drugs with optimal passive absorption characteristics from series of pharmacologically active molecules generated in drug discovery programs.