Solubilization of Liposomes by Weak Electrolyte Drugs. I. Propranolol

The solubilization of dimyristoylphosphatidylcholine (DMPC) liposomes by a weak electrolyte drug, propranolol (PPL) hydrochloride, has been studied as a function of pH, [PPL], [DMPC], and temperature. The solubilization of liposomes at 40°C by 0.2 mM PPL occurred at different rates from 2.9 to 14.4 mM DMPC but converged at complete solubilization after 13 hr at pH 12.0. At the same [PPL], solubilization was complete after 18 days at pH 11.0, but incomplete solubilization occurred at pH 10.0 and not at all at lower pH's. In 14.4 mM DMPC liposomes, solubilization was gradual and proportional to the [PPL] from 0.001 to 0.10 mM up to 95 hr, then rapid thereafter. The [PPL] at which the solubilization efficiency began to increase rapidly was determined to be 0.078 mM. The rate of solubilization was also influenced by the fluidity of the bilayers, a sevenfold increase in the time for complete solubilization being observed upon cooling from 40 to 20°C. Surface tension (st) data confirmed a low critical micelle concentration (CMC) and continued decrease in the st above the CMC. It is concluded that the critical ratio of PPL to DMPC for solubilization occurs in localized regions of the bilayers, with total solubilization at different rates depending on the [PPL] and the physical properties of the liposomes. The processes may be used advantageously to prepare small vesicles or to extract lipids or proteins, more efficiently than detergents, from biological membranes.     

叶酸受体靶向伊马替尼pH敏感脂质体的制备工艺及质量评价研究

摘 要 目的：运用改良的硫酸铵梯度法制备叶酸受体靶向伊马替尼pH敏感脂质体（FSLI）,并评价其质量。方法: 采用改良的硫酸铵梯度法制备FSLI,以包封率为指标进行制备工艺的优化;采用琼脂糖凝胶CL 4B柱分离脂质体和游离药,HPLC法测定FSLI的包封率;运用Zeta PALS粒径电位分析仪测定脂质体的平均粒径及Zeta电位,透射电镜观察脂质体的形态;运用动态透析法考察FSLI在不同pH环境中的药物释放情况,并考察FSLI在不同pH、37℃水浴中的粒径随时间的变化情况。结果: 制得的FSLI为圆形或类圆形的大单室脂质体,平均粒径为（155.2±1.92）nm,Zeta电位为 （29.36±3.21）mV,平均包封率为（90.7±1.70）%。FSLI在pH 7.4的中性环境中,药物释放缓慢,24 h累积释放度为2.76%,而在pH 5.5的酸性环境中,药物释放显著加快,24 h累计释放度均达到93.2%,表现出非常强的pH敏感性能。FSLI在pH 7.4的中性环境中不具有融合性能,而在pH 5.5的酸性环境中,粒径迅速增大,脂质体发生融合。结论:采用改良硫酸铵梯度法制备的FSLI,能获得较高的包封率,并具有良好的pH敏感性能。     

溶液pH对普萘洛尔透皮性能的影响

目的 考察不同pH对普萘洛尔稳定性及离体透皮性能的影响。方法 采用HPLC测定普萘洛尔浓度;以4 500 lx光照及100 ℃高温进行加速试验,研究其在pH5.0～9.0内的稳定性;以正辛醇-磷酸盐缓冲液为模拟系统,采用摇瓶法测定不同pH下的lgP(油水分配系数的对数值);并以大鼠腹部皮肤为模型,采用改良的Franz扩散池考察pH对药物透皮性能的影响。结果 普萘洛尔对热稳定,但在光照条件下发生降解,降解反应符合一级动力学过程,且具有明显的pH依赖性,表现为当pH高于7.4时降解速率常数显著增加。另外,普萘洛尔lgP随pH升高而增大,当pH在7.0以上时,lgP值均大于1,并表现出良好的透皮性能;当pH在7.0以下时,lgP及透皮性能均急剧下降。结论 本研究为普萘洛尔经皮给药制剂的设计与开发提供了实验依据。     

Racial Differences in Drug Response: Isoproterenol Effects Before and After Propranolol

The aim of this study was to determine in young, healthy men the relative contribution of pharmacodynamic factors inherent between two groups known to respond differently to hypertensive therapy. Black (n = 10) and white (n = 10) men received an isoproterenol sensitivity test before and after propranolol (0.1 mg/kg, then 50 µg/min). There were greater increases (twofold) in systolic BP following the 1.0- and 1.5-µg isoproterenol dose (P < 0.05) in the black group. During propranolol there were no differences in free (1)-propranolol concentrations between the groups; however, propranolol decreased resting heart rate in the white group more than in the black group (P < 0.05). Cardiac index decreased less in the black group compared to the white group (P < 0.05). Following the second isoproterenol challenge, there again were greater increases in systolic BP in the black group at both the 10- and the 20-µg isoproterenol dose (P < 0.05). Our study has highlighted the importance of cross-racial studies in evaluating drug effects.     

The Effect of Propranolol on the Electroencephalogram in Normal and Ethanol Dependent Rats

Abstract: Electroencephalograms were recorded from scalp electrodes placed over the cerebral hemispheres of rats withdrawing from ethanol. The purpose was to investigate whether administration of the β-adrenergic receptor blocker propranolol changed the EEG during that condition. The animals were artificially ventilated with an air mixture of 70% N₂O:30% O₂ and PaO₂ PaCO₂ and mean arterial blood pressure were kept constant. There were no significant differences in EEG frequencies or amplitudes between abstinent animals and controls neither before nor 20 min. after intravenous administration of propranolol 2 mg/kg body weight. Similarly, there was no EEG changes during 3 min. of photic stimulation. It has been reported that electrophysiological abnormalities during physical ethanol dependence are generated in subcortical brain structures but the 4 day intoxication period used in the present study seems too short to involve the cerebral cortex.     

尿中普萘洛尔对映体葡醛酸苷的HPLC测定

建立了人尿中普萘洛尔对映体葡醛酸苷无需水解、直接测定的HPLC法.采用生物合成获得普萘洛尔葡醛酸苷,并经C18固-液萃取纯化、浓集,作为对照品储备液.采用荧光检测器,激发波长和发射波长分别为310和339nm.线性范围0.0232～5.8μmol/L,r=0.9999.平均回收率为99.8%±5.01%,日内和日间精密度分别小于1.08%和2.86%.     

Liposomes in the Treatment of Infections

Abstract

The use of liposomes in the treatment of severe infections is under investigation. Classical liposomes which localize in cells of the mononuclear phagocyte system (MPS) can be exploited in two ways. First for targeting of macrophage modulators such as muramyl peptides or IFN-γ, to stimulate the cells of the MPS to maximal blood clearance capacity. This enhanced nonspecific anti-infectious resistance is important as in immunocompromised patients micro-organisms frequently appear in the blood from a local infection. Secondly, classical liposomes are successfully used as carriers of antibiotics in experimental intracellular parasitic-, viral-, fungal- or bacterial infections in MPS tissues. Based on these data extensive studies in patients with severe fungal infections have demonstrated successful treatment with liposomal or lipid-complexed amphotericin B. More recently, liposomal amphotericin B appeared to be effective in patients with drug-resistant visceral leishmaniasis. For the treatment of Mycobacterium avium complex infection in AIDS patients the efficacy of liposomal gentamicin is under investigation. With respect to infections in non-MPS tissues the applicability of Stealth® liposomes characterized by long circulation half-lives is under investigation. Substantial localization of these liposomes in infected lung tissue of rats was demonstrated. Preliminary data in experimental bacterial lung infection showed superior efficacy of antibiotic encapsulated in Stealth® liposomes.     

Effect of Neutral Liposomes on Corneal and Conjunctival Transport of Didanosine

The objective of this study is to determine the effect of various neutral liposomes on corneal and conjunctival permeability of didanosine (ddI), an antiviral drug. Multi-lamellar vesicles (MLVs), large unilamellar vesicles (LUVs), and sonicated multi-lamellar vesicles (SMLVs) encapsulating ddI (with trace quantities of 3HddI) were prepared using distearoyl phosphatidylcholine (DSPC), a neutral lipid. The liposomes contained 14C-cholesteryl oleate as a lipid tracer. Liposome formulations containing free and encapsulated drug (f + e) and those containing only encapsulated drug (e) of an equal quantity were compared with free drug in this study. The permeability studies were conducted in the mucosal to serosal direction across excised rabbit cornea and conjunctiva. The percent encapsulation of ddI in MLVs, LUVs, and SMLVs was 25.66 0.30, 26.56 0.57, and 19.41 0.30, respectively. The mean particle size of MLVs, LUVs, and SMLVs containingfree and encapsulated drug was 3058, 774, and 270 nm, respectively. With all liposome formulations tested, the percent uptake of lipid by tissues was higher compared to ddI uptake. While ddI permeated across the tissues, the lipid tracer did not permeate in detectable quantities.The SMLV(e) formulation was better than the SMLV(f + e) formulation with respect to initial flux and tissue uptake in both tissues and permeability across conjunctiva. In general, the permeability coefficient, initial flux, and tissue levels of ddI at the end of the transport study were lower in the presence of all liposome formulations compared to free drug. Thus, neutral liposomal encapsulation is not a suitable approach to enhance the corneal or conjunctival transport or uptake of ddI.     

Towards the Predictability of Drug-Lipid Membrane Interactions: The pH-Dependent Affinity of Propranolol to Phosphatidylinositol Containing Liposomes

Purpose. Prediction of the pH-dependent affinity of (RS)-[³H]propranolol to mixed phosphatidylcholine (PhC)/phosphatidylinositol(Phl) membranes from the partitioning in the single lipid liposome/buffer systems. Methods. Partition studies in liposome/buffer systems were performed by means of equilibrium dialysis at 37°C between pH 2 and 11 at a molar propranolol to lipid ratio of 10^–6 to 10^–5 in the membrane. Results. The Phl membrane more strongly attracts the protonated (RS)-[³H]propranolol than the neutral solute, i.e. the partition coefficient of the protonated base (P_i) is 17430 ± 1320, P of the neutral compound (P_n) is 3110 ± 1650. In the PhC-liposome system P_i is 580 ± 17, P_n 1860 ± 20. The partition coefficients show an exponential dependence on the molar Phl fraction in mixed liposomes. The partitioning in mixed PhC/Phl membranes is predictable from P_n and P_i in the single lipid liposome systems. Conclusions. The negative charge of biological lipid membranes causes strong electrostatic interactions with positively charged solutes. This strong attraction is not predictable from the octanol/buffer partition system, but it is important regarding drug accumulation in the tissue and drug attraction by certain lipids in the vicinity of membrane proteins.     

川芎嗪脂质体对人白血病细胞株K562多药耐药逆转作用的研究

目的:研究川芎嗪脂质体在体外对肿瘤细胞的细胞毒作用和对多药耐药的逆转作用,并研究脂质体剂型对上述作用的影响.方法:MTT法检测川芎嗪脂质体以及川芎嗪原料对照组、空白脂质体对照组对耐阿霉素的人白血病细胞株(K562/ADM)的细胞毒性和多药耐药逆转作用.结果:川芎嗪脂质体对K562/ADM有明显的细胞毒作用.非细胞毒性剂量(生长率≥95%)为2.0μg·ml-1,该浓度下可显著降低ADM对K562/ADM细胞的IC50(P＜0.01),抗药性逆转为11.2倍.低毒剂量(生长率85～90%)为2.6μg·ml-1,抗药性逆转为178倍.2.0μg·ml-1的川芎嗪原料对照组可显著降低ADM对K562/ADM细胞的IC50(P＜0.01),抗药性逆转为7.9倍;同比稀释的空白脂质体对照组可显著降低ADM对K562/ADM细胞的IC50(P＜0.01),抗药性逆转为4.9倍.结论:川芎嗪脂质体体外对人白血病细胞K562/ADM有明显的细胞毒作用和MDR逆转作用;在一定浓度范围内川芎嗪脂质体的细胞毒性与逆转作用有剂量依赖性;脂质体本身对逆转有增效作用.     

基于pH梯度法的盐酸普萘洛尔立方液晶纳米粒的制备

目的 制备具有较高包封率的盐酸普萘洛尔立方液晶纳米粒（PPL- Cubs）。方法 采用pH梯度法制备PPL-Cubs;以粒径、多分散指数为评价指标,优化空白立方液晶纳米粒（B-Cubs）制备的高压均质压力、高压均质次数、单油酸甘油酯用量及泊洛沙姆407用量;以包封率等为评价指标,优化外水相pH值、内水相pH值、载体/药物比、载药温度、载药时间、B-Cubs粒径和多分散指数、药物浓度等。结果 高压均质压力为900 bar、均质次数为7次、单油酸甘油酯用量为25%、泊洛沙姆407用量为5%时,制得的B-Cubs具有较小的粒径和多分散指数。外水相pH值为8.5、内水相pH值为3.0、载体/药物比为6∶1、载药温度为20 ℃、载药时间为15 min、药物浓度为1%时,制得的PPL-Cubs包封率较高;B-Cubs粒径和多分散指数对制得的PPL-Cubs包封率无明显影响。结论 pH梯度法能制得较高包封率的盐酸普萘洛尔立方液晶纳米粒。     

紫杉醇脂质体制备及体外抑制两种细胞的作用

目的:制备紫杉醇脂质体并考察体外对人卵巢癌上皮细胞(SKOV-3)和人脐静脉内皮细胞(HUVEC)两种细胞的抑制作用。方法:采用薄膜分散法制备紫杉醇脂质体,透射电镜观察脂质体的形貌,激光粒度仪测量粒径和Zeta电位,MTT法检测脂质体体外对SKOV-3和HUVEC的抑制作用。结果:制备的脂质体呈圆形或类圆形,平均粒径为54.6 nm,Zeta电位为-40.9 mv,体外释放符合Higuchi方程。体外细胞结果实验显示,紫杉醇能同时抑制肿瘤细胞和血管内皮细胞的生长。结论:紫杉醇脂质体较紫杉醇注射液(TAXOL)能更有效的抑制SKOV-3和HUVEC的生长。     

pH梯度法制备阿霉素脂质体

采用ｐＨ梯度法制备了阿霉素脂质体．用ＳｅｐｈａｄｅｘＧ－２５微型柱分离－紫外分光光度法测定阿霉素脂质体包封率．结果表明，当药物／磷脂重量比为０．０２～０．０５时，包封率均大于９０％；体外泄漏研究显示，２５℃时，２ｈ内泄漏百分率约为２．７％，保证了临床应用．     

季铵化程度对N-三甲基壳聚糖包衣脂质体生物安全性的影响

目的:研究N-三甲基壳聚糖(TMC)的季铵化程度(DQ)对TMC包衣脂质体安全性的影响规律,为其作为药物载体的安全应用提供实验依据.方法:分别制备未包衣的脂质体及用不同DQ的TMC(TMC20,TMC40及TMC60)包衣的脂质体;采用静脉注射及腹腔注射,对其小鼠急性毒性进行考察,分别得到静脉注射及腹腔注射的LD50;对其注射刺激性、过敏性及溶血性进行考察.结果:未包衣脂质体及TMC20,TMC40、TMC60包衣脂质体静脉注射的LD50及95%置信限分别为808.1(676.4～1 009.5)、721.9(584.0～971.9)、319.1(261.9～399.0)、224.1(178.8～302.6)mg·kg-1;未包衣脂质体及各种包衣脂质体腹腔注射的LD50及95%置信限分别为1 158.3(958.4～1 443.0)、1 109.5(899.9～1 415.4)、573.7(480.2～716.6)、342.3(269.7～534.2)mg·kg-1.与未包衣的脂质体相比,TMC20包衣后小鼠静脉注射及腹腔注射的LD50无明显变化(P>0.05),而TMC40及TMC60包衣脂质体的相应LD50有较明显的降低(P<0.05);包衣脂质体间相比较,相应LD50随着DQ的增加而逐渐下降(P<0.05);在实验剂量时,各组脂质体均无注射刺激性、过敏性及溶血性.结论:TMC的DQ对其包衣的脂质体生物安全性有较明显的影响,DQ越大,LD50越低.     

Effect of Receiver Fluid pH on in Vitro Skin Flux of Weakly lonizable Drugs

In in vitro skin permeation experiments, the pH of viable epidermis is readily conditioned by the receiver fluid. For weakly ionizable compounds, the flux determined experimentally thus depends on the receiver fluid pH. The purpose of the present work is to characterize this pH effect, since nonphysiological conditions have often been used in the receiver fluid to enhance the solubility of the subject compounds. A transport model was developed to analyze the above-mentioned pH effect of the receiver fluid on the steady state flux of weakly ionizable drugs. The results showed that the skin flux had a strong dependence on pH for those compounds with high intrinsic partition coefficients. Experimentally, this pH effect was observed with a model acid and a model base. The skin flux was found to have a profound dependence on the receiver fluid pH. This dependence also correlates with the octanol/water partition coefficient of the molecule. It was concluded that the use of a physiological receiver fluid would be crucial for a realistic estimation of transdermal potential. The results also suggested that, for weakly ionizable compounds with high partition coefficients, the viable epidermis could be a significant transport barrier for systemic absorption.     

Sustained Propranolol Delivery and Increased Oral Bioavailability in Dogs Given a Propranolol Laurate Salt

Pharmaceutical Research -     

主动载药法制备两亲性药物脂质体的研究进展

脂质体跨膜梯度主动载药方法主要有pH梯度法和硫酸铵梯度法.本文介绍了两亲性药物脂质体跨膜梯度载药方法的原理、制备工艺、影响包封率的因素等国内外研究进展.     

Study on Liposomes of Mimetic Red Cell

本项研究从仿红细胞出发，将增加双分子脂膜亲水性的ＰＥＧＰＥ（聚乙二醇与磷脂酰乙醇胺的衍生物）及增加双分子脂膜硬度的ＳＭ（神经鞘磷脂）掺入脂质体，比较其在体外稳定性、体内分布及清除半衰期。结果证明该类脂质体稳定性提高，体内被肝、脾清除减少，血循环中半衰期显著延长。本研究为脂质体载药至非网状内皮系统的靶向给药打下基础。     

甘草次酸修饰多西紫杉醇脂质体的制备和体外抑瘤效果

目的:制备甘草次酸(GA)修饰的多西紫杉醇脂质体,并初步考察其体外抗肿瘤效果.方法:化学合成甘珀酸十八醇酯(18-GA-Suc)作为修饰材料,采用薄膜分散法制备甘草次酸修饰的多西紫杉醇脂质体,考察影响脂质体包封率的因素.采用MTT法评价脂质体对HepG2细胞的体外抑瘤效果.结果:18-GA-Suc修饰的DX脂质体的体外抑瘤效果强于未修饰的DX脂质体,并且抑瘤效果随着载体中18-GA-Suc的增加而增强.结论:甘草次酸修饰的脂质体有望成为新型肝靶向的抗肿瘤载体.     

Comparison of the Effects of Dimyristoyl and Soya Phosphatidylcholine Liposomes on Human Fibroblasts

The in vitro effects of prolonged exposure (8 days) of human skin fibroblasts to several concentrations of extruded dimyristoyl (dm-PC) and soya phosphatidylcholine (soya-PC) liposomes were compared. Prepared liposome suspensions were added to the fibroblast culture medium at phospholipid concentrations of 10, 50, 100, 200, and 300 mu M. Survival curves and values of 50% inhibitory concentration (IC₅₀) and area under the curve (AUC) were used to compare the response of the fibroblasts to the two types of liposomes. The effect of the incorporation of vitamin E in the liposomal preparations also was determined. Fibroblasts showed greater sensitivity toward the soya-PC liposomes (IC₅₀ = 150 mu M) than the dm-PC liposomes (IC₅₀ = 212 mu M). The presence of vitamin E in the soya-PC liposomes led to a 1.9-fold increase in the IC₅₀, while dm-PC liposomes containing vitamin E showed an IC₅₀ that was 1.1 times higher than that shown by control vitamin-free liposomes. Soya-PC liposomes containing vitamin E at a molar ratio of 10:0.5 (phospholipid:vitamin)were best tolerated by the fibroblasts (IC₅₀ &gt; 300 mu M). It would appear that dm-PC liposomes are better tolerated by fibroblasts than those composed of soya-PC. However, the incorporation of vitamin E into the liposomes seems to reverse this effect, and it is the vitamin-containing soya-PC liposomes that are most compatible with the growth of fibroblasts in culture.