白蛋白微球的制备方法及质量评价的研究进展

微球给药系统在实现药物靶向给药,其在缓、控释放等方面表现出良好的应用前景,因而成为近年来药剂学领域的研究热点之一。本文综述了白蛋白微球的主要制备方法及其优缺点,并对白蛋白微球的质量评价方法进行归纳。     

盐酸克仑特罗双层缓释片的研制及体内外评价

目的研制盐酸克仑特罗缓释双层片并考察其体外释放及犬体内药动学。方法以羟丙甲纤维素和乙基纤维素为骨架材料,分别采用粉末直接压片和湿法制粒技术制备盐酸克仑特罗双层缓释片。测定其体外释放度和Beagle犬口服单剂量盐酸克仑特罗普通片和双层缓释片后血浆中药物的浓度,推算药动学参数。结果盐酸克仑特罗双层缓释片体外释放符合Higuehi方程。盐酸克仑特罗双层缓释片和普通片的有关药动学参数如下：t1/2分别为（11．90±1．42）,（6．18±0．81）h;Cmax为（5．86±0．33）,（11．43±0．64）ng·mL^-1;tmax为（3．17±0．73）,（2．02±0．35）h;AUC0～1．为（65．08±2．63）,（68．00±2．89）ng·h·mL^-1。结论盐酸克仑特罗双层缓释片体外具有明显的速释和缓释特性。体内平均滞留时间长于普通片。体外释放和体内吸收有良好的相关性。     

心脏靶向超声造影剂空气白蛋白微球的研究(1)

按正交设计筛选了用发射超声能量合成白蛋白空气微球的较佳制备工艺,并对微球的形态、粒径与粒径分布、稳定性、安全性和显影效果进行了系统研究。结果表明：微球的平均直径为 ４．２±０． ２ μｍ,平均浓度为（３．５±１．２）×１０８／ｍｌ。９２．５％的微球直径小于１０μｍ,微球在２℃～５℃条件下能稳定储存１２个月,经兔外用静脉注射后能顺利通过肺微循环到达左心室致使左心室靶向超声显影。     

白蛋白微球的制备及影响粒径大小的因素

介绍了聚合物分散法制备白蛋白微球的方法。重点研究聚甲基丙烯酸甲酯(PMMA)溶液的粘度和乳化过程中的搅拌速度对微球粒径大小的影响。结果表明,PMMA溶液的粘度越大,乳化时搅拌速度越快,微球的粒径分布越小。     

Albumin Microspheres as an Ocular Delivery System for Pilocarpine Nitrate

Pilocarpine nitrate loaded egg albumin microspheres were prepared by thermal denaturation process in the size range of 1-12 μm. A series of batches were prepared to study factors, which may affect the size and entrapment efficiency of drug in microspheres and optimized the process. Drug loaded microspheres so obtained were evaluated for their size, entrapment efficiency, release rate and biological response. Electron photomicrographs were taken (8000X) to study the morphological characteristics of microspheres. The entrapment and encapsulation of pilocarpine after process optimization was found to be 82.63% and 62.5% respectively. In vitro dissolution rate studies revealed that the release of drug from the microspheres followed spherical matrix mechanism. Biological response of microspheric suspension was measured by reduction in intraocular pressure in albino rabbit eyes and compared with marketed eye drops. Various pharmacokinetic parameters viz. onset of action, duration of action, Tmax and AUC were studied. A measurable difference was found in the mean miotic response, duration and AUC of pilocarpine nitrate microspheric suspension.     

多柔比星白蛋白微球的生物性质及其在大鼠体内的分布

目的:研究多柔比星(DR)白蛋白微球生物性质及其在大鼠体内的分布。方法:采用胰酶降解法研究微球的生物降解性能并确定交联剂浓度及固化时间与其降解时间的关系;应用透析法研究DR白蛋白微球和DR水溶液的体外溶出/释药性;测定大鼠尾静脉分别注射DR白蛋白微球和DR水溶液2.5 mg·kg~(-1)后体内各组织中分布浓度。结果:降解时间随交联剂浓度和交联时间增加而延长,固化时间以90 min为宜;DR水溶液在6 h内溶出完全,微球混悬液在168 h释放量仅达80%;微球组的DR在肝和脾中分布浓度显著高于水溶液组(P<0.01),其它组织中则低于水溶液组。结论:DR白蛋白微球具有明显的缓释作用,对肝和脾表现出显著靶向性,对心脏具有很低的亲和性。     

聚乙二醇对超声造影剂空气白蛋白微球制备性质的影响

目的:研究加入聚乙二醇(PEG)对制备空气白蛋白微球的影响.方法:在制备空气白蛋白微球过程中,将其处方中加入PEG,并测定制得微球的浓度及粒径.结果:加入PEG后,测得3批样品的平均微球浓度为3.50×108个/mL,平均直径为4.27μm;相应的不加PEG的样品分别为2.70×108个/mL和4.14μm.结论:可在制备空气白蛋白微球时加入聚乙二醇,这为在微球上联接配体等进一步研究奠定了基础.     

Release of Human Serum Albumin from Poly(lactide-co-glycolide) Microspheres

Human serum albumin (HSA) was encapsulated in a 50:50 copolymer of DL-lactide/glycolide in the form of microspheres. These microspheres were used as a model formulation to study the feasibility of controlling the release of large proteins over a 20- to 30-day period. We show that HSA can be successfully incorporated into microspheres and released intact from these microspheres into various buffer systems at 37°C. A continuous release of the protein could be achieved in physiological buffers at 37°C over a 20- to 30-day period from microspheres with high protein loadings (11.6%). These results demonstrate the potential of poly(DL-lactide-co-glycolide) microspheres for continuous delivery of large proteins.     

沙美特罗白蛋白微球在大鼠体内的药动学

目的:研究沙美特罗白蛋白微球在大鼠体内的药动学特征.方法:12只大鼠随机分成沙美特罗白蛋白微球组和沙美特罗溶液组,每组6只,分别灌胃给药(20 μg·kg -1)后不同时间取血,HPLC法测定血药浓度,用3P97软件计算药动学参数.结果:沙美特罗溶液和沙美特罗白蛋白微球在大鼠体内的主要药动学参数:tmax分别为(0.73±0.15)和(3.22±0.20)h,Cmax分别为(1.10±0.19)和(1.25±0.24) μg·ml-1,t1/2ka分别为(0.35±0.05)和(1.58±0.07)h,t1/2ke分别为(2.30±0.85)和(8.21±1.20)h,AUCo→∞分别为(1.55±0.25)和(3.45±0.55) mg·h·L-1.结论:与沙美特罗溶液相比,沙美特罗白蛋白微球具有明显的缓释效果,同时提高了药物的生物利用度.     

含有甲氨蝶呤（MTX）的牛血清白蛋白（BSA）微球——Ⅰ.制剂学部分

用乳化技术制备了三种微球体系,分别命名为Ⅰ、Ⅱ、Ⅲ。Ⅰ是乳剂通过戊二醛交联的方法制成,Ⅱ是在戊二醛交联的基础上用甘氨酸处理的产品,Ⅲ是乳剂通过加热固化法而制得的微球。结果发现,相同的乳剂经交联法制得的微球其直径比加热固化法制得的微球大。微球Ⅱ和微球Ⅰ有更好的亲水性。微球Ⅰ和Ⅱ中 MTX 的含量分别为2.73±0.05%和2.87±0.12%。Ⅲ是空白微球表面吸附了 MTX 结晶的一种体系。微球中 MTX 的体外释放实验证实,Ⅰ和Ⅱ在 pH 7.4的磷酸盐缓冲液中,药物释放达24小时之久其机理属 Higuchi 扩散。     

苦参碱白蛋白微球的制备及性质

以乳化-化学交联法制备苦参碱白蛋白微球。采用扫描电镜、差示扫描量热法和X-射线衍射法考察微球的特性,并用动态透析法考察体外释药规律。所得微球外观圆整,包封率为(83.55±3.17)%,载药量为(6.19±0.24)%,平均粒径(0.94±0.15)μm。体外释放试验结果显示,苦参碱白蛋白微球具有明显的缓释作用。     

肺靶向顺铂磁性白蛋白微球的制备及性质

采用乳化复合技术制备粒径1.25～3um的顺铂磁性白蛋白微球。该微球呈圆球形,大小分布均匀,在0.05T的弱磁场中具有磁响应性,在水中分散性好。载药量为8.8%(mg/mg）,包封率为96.21%,临界相对湿度（CRH)=73.3%。含药微球在体外释药较快,缓释作用不明显。体内外磁定位实验表明,微球可浓集于靶区。该制剂在室温下放置3个月,性质确定。     

沙美特罗白蛋白微球的制备与体外释放度考察

目的:制备沙美特罗白蛋白微球,并考察其体外释放性能。方法:以白蛋白为载体,采用乳化交联法制备沙美特罗白蛋白微球。在单因素考察的基础上,利用正交设计优化沙美特罗白蛋白微球制备工艺,并对微球的粒径,形态,体外释放特性进行研究。结果:制得的微球形态圆整,平均粒径为（16.82±1.25）μm,平均载药量为（52.08±3.26）%,平均包封率为（60.54±3.17）%,体外释放符合Higuchi方程,Q=11.5846t_1/2-1.207（r=0.9985）。结论:沙美特罗白蛋白微球体外释放特性符合长效制剂特征。     

3种方法分析白蛋白空气微球的球壁蛋白

采用高效液相色谱（HPLC）、醋酸纤维膜电泳和半微量定氮法测定5％声振人血白蛋白空气微球制剂中球壁蛋白的含量。HPLC确证了球壁蛋白主要成分为人血白蛋白；醋酸纤维膜电泳测定空气人血白蛋白微球制剂中人血白蛋白纯度＞90％；半微量定氮法测出球壁蛋白含量为3．56％左右，99％正常值范围为2．52％～4．60％。     

Albumin Microspheres as a Drug Delivery System: Relation Among Turbidity Ratio,Degree of Cross-linking,and Drug Release

The degree of cross-linking of albumin microspheres, with and without drug, was assessed using turbidity measurements carried out in the presence of water and the protein denaturant guanidine hydrochloride (GuHCl) at a concentration that disrupted noncovalent bonds while having no effect on covalent bonds. The measurements allowed calculation of a turbidity ratio (T _G/T _W), expressed as the ratio of the turbidity of albumin microspheres in 6 M GuHCl (T _G) divided by that in water (T _w). A linear relation existed between T _G/T _W and the (i) temperature at which the microspheres were prepared, (ii) concentration of the cross-linking agent glutaraldehyde, and (iii) time of exposure to a second cross-linking agent, formaldehyde vapor, three conditions that increase the degree of cross-linking. The turbidity ratio also increased as the concentration of the albumin solution used to prepare the microspheres increased from 25 to 50%. Drug release from the microspheres consisted of an initial, rapid, burst followed by a second, slower, phase. The rates in both release phases were inversely related to the turbidity ratio, suggesting that this parameter has utility as an indicator of the degree of cross-linking in albumin microspheres.     

去甲斑蝥素白蛋白微球在大鼠体内的药动学及组织分布研究

目的：研究去甲斑蝥素（NCTD）白蛋白微球在大鼠体内的药动学及组织分布情况。方法：以市售去甲斑蝥素注射液为参比,测定NCTD白蛋白微球在大鼠体内血浆中的药动学模型及药动学参数,用靶向效率来评价其在大鼠体内的组织分布及靶向性。结果：NCTD白蛋白微球在大鼠体内的药动学模型符合二室模型,主要药动学参数为：t_（1/2α）=（0.57±0.11）h,t1/2β=（20.69±1.06）h,CL=（0.016±0.002 4）ml·h·kg^-1,AUC0～∞=（63.41±9.08）mg·h·L^-1,其在肝脏,脾脏,心脏及肾脏的靶向效率分别为为2.36,1.78,0.43和0.35。结论：与去甲斑蝥素注射液相比,NCTD白蛋白微球能提高对肝脏,脾脏的趋向性,减少对肾脏及心脏的分布,有利于提高其治疗作用,减少不良反应。     

Adsorption of Salmon Calcitonin to PLGA Microspheres

Purpose. The interaction of salmon calcitonin (sCT) and poly (d,l-lactide-co-glycolide) was detected during preparation and evaluation of microspheres. The purpose of this study was to quantitate the extent and nature of the interaction. Methods. Blank microspheres were prepared by an aqueous emulsification solvent extraction technique. Adsorption studies were carried out at six concentrations of sCT and three concentrations of microspheres. Adsorption isotherms were constructed using the Langmuir and Freundlich treatments. Results. Adsorption at 1 mg/ml sCT concentration resulted in almost complete depletion of the peptide from the adsorption medium with the time to reach maximum adsorption decreasing with increasing microsphere concentration. At sCT concentrations below 100 µg/ml, a true equilibrium occurred in 1 hour or less while at higher concentrations (up to 350 µg/ml), a transient equilibrium was reached in 1 to 2 hours, followed by further adsorption of the peptide. The adsorption followed the Langmuir isotherm at concentrations below 200 µg/ml, indicating formation of a monolayer. Multilayer interaction, described by the Freundlich isotherm, occurred at higher concentrations and resulted in complete depletion of sCT from the adsorption medium. The affinity constant during monolayer formation was 0.09 and the plateau surface concentration was 5.1 µg/mg. The multilayer peptide-peptide adsorption showed a lower affinity (0.025) but higher capacity (24 µg/mg) than the monolayer peptide-polymer adsorption. Conclusions. The results show that poly (d,l-lactide-co-glycolide) microspheres have a high adsorption capacity for sCT which must be considered in formulating a controlled delivery product of this peptide.     

Determination of Air Content in Protein Microspheres

Pharmaceutical Research -     

加替沙星与牛血清白蛋白相互作用的研究

目的研究不同酸度条件下, 加替沙星与牛血清白蛋白之间的相互作用.方法采用荧光光谱和紫外光谱法进行研究.结果运用荧光猝灭双倒数图计算了在不同条件下二者的结合常数K, 根据Foster非辐射理论计算在正常生理条件下二者的结合距离r, 并通过热力学参数确定了二者的作用力类型.结论加替沙星与牛血清白蛋白之间有较强的相互作用, 以电荷作用力为主.     

Stabilization and Controlled Release of Bovine Serum Albumin Encapsulated in Poly(D,L-lactide) and Poly(ethylene glycol) Microsphere Blends

Purpose. The acidic microclimate in poly(D, L-lactide-co-glycolide) 50/50 microspheres has been previously demonstrated by our group as the primary instability source of encapsulated bovine serum albumin (BSA). The objectives of this study were to stabilize the encapsulated model protein, BSA, and to achieve continuous protein release by using a blend of: slowly degrading poly(D, L-lactide) (PLA), to reduce the production of acidic species during BSA release; and pore-forming poly(ethylene glycol) (PEG), to increase diffusion of BSA and polymer degradation products out of the polymer. Methods. Microspheres were formulated from blends of PLA (Mw 145,000) and PEG (Mw 10,000 or 35,000) by using an anhydrous oil-in-oil emulsion and solvent extraction (O/O) method. The polymer blend composition and phase miscibility were examined by FT-IR and DSC, respectively. Microsphere surface morphology, water uptake, and BSA release kinetics were also investigated. The stability of BSA encapsulated in microspheres was examined by losses in protein solubility, SDS-PAGE, IEF, CD, and fluorescence spectroscopy. Results. PEG was successfully incorporated in PLA microspheres and shown to possess partial miscibility with PLA. A protein loading level of 5% (w/w) was attained in PLA/PEG microspheres with a mean diameter of approximately 100 m. When PEG content was less than 20% in the blend, incomplete release of BSA was observed with the formation of insoluble, and primarily non-covalent aggregates. When 20%-30% PEG was incorporated in the blend formulation, in vitro continuous protein release over 29 days was exhibited. Unreleased BSA in these formulations was water-soluble and structurally intact. Conclusions. Stabilization and controlled relaease of BSA from PLA/PEG microspheres was achieved due to low acid and high water content in the blend formulation.