pH-Sensitive Self-Assembled Microspheres Composed of Poly(Ethyleneimine) and Cinnamic Acid

Microspheres which were sensitive to pH change were developed by utilizing cinnamic acid (CA) as a physical cross-linker for poly(ethyleneimine) (PEI). At pH 7.0, the microspheres were efficiently formed at the PEI/CA ratio of 1:3.4, 1:5.1, and 1:7.1 (w/w), which corresponded to the protonated amino group/deprotonated carboxyl group ratio of 5:5, 4:6, and 3:7. The mean diameter of wet microspheres was 3.2 ± 0.3 to 8.8 ± 0.5 μm and that of dry ones was 1.7 ± 0.2 to 2.7 ± 0.2 μm. The microspheres were disappeared upon the alkalification, possibly because the electrostatic interaction between PEI and CA was slackened down and the hydrophobic interaction among CA molecules was weakened. At pH 5.0 and 7.0, the microsphere released its content in a sustained manner and the release degree in 24 h was less than 40%. Whereas, at pH 8.0 and 9.0, the microsphere exhibited a burst release and the release degree in 24 h was greater than 80%. In the alkali condition, not only the electrostatic interaction between PEI and CA but also the hydrophobic interaction among CA molecules became weaker, leading to the disintegration of the microsphere and resulting in a burst and intensive release.     

生物降解聚酯包埋利福平缓释微球的制备及释放行为

以生物可降解乙交酯和丙交酯的无规共聚物(PLGA)为载体,将抗结核病药利福平溶解于PLGA的有机溶液中,采用通常乳化-溶剂挥发方法制备了药物缓释微球.研究了影响微球制备的工艺条件.用电子显微镜观察了微球及降解后的表面形态,测定了微球粒径及载药量,评价了载药微球的体外释放行为.结果表明,以质量分数为1%的明胶为稳定剂,制备的微球形态完整,粒径范围为10～30μm,微球中利福平的平均质量分数为24.3%.体外释药时间可以通过高分子的降解速率来调控,本实验的释药时间可以在42～84d之间调控,药物缓释达到了理想的零级动力学释放.因此,利福平PLGA微球具有显著的长效、恒量药物缓释作用.     

A dual microsphere based on PLGA and chitosan for delivering the oligopeptide derived from BMP-2

In this paper, we document the process and findings of preparing dual poly (lactide-co-glycolide)/chitosan microspheres (PLGA/CS MSs) for osteoinductive oligopeptide derived from BMP-2 (abbreviated as Peptide-24). Through adjusting the amount of Peptide-24, three kinds of PLGA/CS MSs were successfully constructed in twice encapsulations. We studied the morphology, size distribution and loading efficiency of the PLGA/CS MSs. We also focused on the pH change of the environment and the molecular weight of the matrix during the degradation process of PLGA/CS MSs. More specifically, the release of Peptide-24 from three kinds of PLGA/CS MSs was monitored in PBS at 37 °C and pH 7.4. The structural stability of the released Peptide-24 was detected by Far-UV circular dichroism and MALDI-TOF-MS analysis. The mean sizes of the three kinds of PLGA/CS MSs are 47.5, 63.0 and 89.1 μm; and their drug-loading rates are 2.61, 3.21 and 2.21%, respectively. Comparing with Chitosan microspheres (abbreviated as CS MSs), the PLGA/CS MSs have excellent release curves with zero-order kinetics and controllable model. The incubation solution of PLGA/CS MSs avoided producing acid environment as poly (lactide-co-glycolide) microspheres (PLGA MSs) did, which was explained by analyzing the molecular weight of the matrix. The released oligopeptide kept its original structure and relative molecular weight throughout the procedures of encapsulation, storage and release. This indicates its structure stability. Thus, we conclude that dual PLGA/CS MSs is a promising vehicle that is suitable for the delivery of bioactive factors.     

载有胰岛素的可生物降解微球的制备与表征

用乙交酯与丙交酯的无规共聚物(PLGA)和聚乙二醇单甲醚-聚丙交酯两嵌段共聚物(MPEG-PLA)的合金作为囊材料,包裹胰岛素固体粉末,包裹率分析表明,固体粉末法对胰岛素的包裹率高于双乳液法.所得微球球形很好,尺寸在1～3μm范围,剖面具有核壳结构,胰岛素以晶粒的形式包裹在高分子壳层中.两种高分子在凝聚过程中发生相分离,在壳层中有分层现象.测定微球的体外释放行为,由聚合物合金制备的微球的暴释现象得到了缓解,两种聚合物的配比不同,其暴释缓解的程度也不一样.     

肾上腺髓质素微球复合PLGA/纳米羟基磷灰石支架材料的制备及生物活性评价

利用离子乳化交联法制备了负载肾上腺髓质素的壳聚糖微球,应用热致相分离法制备了乳酸和乙醇酸共聚物/纳米羟基磷灰石(PLGA/nHA)支架材料并在其中包覆载药微球.通过扫描电子显微镜、体外释放行为、材料溶血行为、碱性磷酸酶(ALP)活性的测定、支架材料表面细胞荧光染色和MTT[3-(4,5-二甲基噻唑-2)-2,5-二苯基四氮唑溴盐]比色法等手段综合评价载药支架材料的性能及生物活性.结果表明,微球直径均匀,载药支架孔径大小合适并相互穿通.支架材料的溶血率小于5%,符合医用材料的溶血实验要求.载药支架及支架材料本身对成骨细胞及血管内皮细胞的增殖以及成骨细胞的分化均有一定的促进作用.     

Preparation and properties of monodispersed rifampicin-loaded poly(lactide-co-glycolide) microspheres

Monodispersed rifampicin (RFP)-loaded poly(lactide-co-glycolide) (PLGA) microspheres were prepared by a solvent evaporation method. In order to control the sizes of the microspheres, a membrane emulsification technique using Shirasu porous glass (SPG) membranes was applied. RFP/PLGA microspheres with the average diameters of 1.3, 2.2, 5.2, and 9.0 microm were obtained. They were relatively monodisperse and the values of the coefficient of variation (CV) for the size distributions of the microspheres were in the range between 7.0 and 16.0%. The loading efficiency of RFP was in the range between 50.3 and 67.4% independent of the microsphere size. The release ratio of RFP from RFP/PLGA microspheres was measured in pH 7.4 PBS at 37 degrees C. From RFP/PLGA microspheres with average diameters of 1.3 and 2.2 microm, almost 60% of RFP loaded in the microspheres was released in the initial day and the release was terminated almost within 10 days. On the other hand, from those with average diameters of 5.2, and 9.0 microm, the release of RFP was observed even 20 days after the release started.     

Poly(lactic-co-glycolic acid) microspheres for the controlled release of huperzine A: in vitro and in vivo studies and the application in the treatment of the impaired memory of mice

Huperzine A loaded poly(D,L-lactic-co-glycolic acid) (PLGA) microspheres were prepared by an oil/water (o/w) solvent evaporation technique. With a decrease of the ratio of o/w from 1 : 100 to 1 : 50, the encapsulation efficiency was reduced about 4%. Increasing the PVA concentration from 0.5 to 2% reduced the percentage encapsulation efficiency of huperzine A from 60.7 to 47.4% and the particle size of microspheres from 84.2 to 26.2 microm. The addition of stearic acid improved the encapsulation efficiency, but also accelerated the in vitro release of hupezine A from microspheres. After i.m. administration of huperzine A loaded microspheres in mice, huperzine A was sustained released from the PLGA microspheres up to 12 d with a low initial burst. Passive avoidance test of mice showed that the microspheres formulation offered an improved therapeutic efficiency in the treatment of the impaired memory of the mice superior to injection gastric (i.g.) administration of huperzine A suspension at the same dose, whose therapeutic efficiency was similar as that of a 50% reduced dose of the microspheres formulation.     

以PHBV-PLGA为载体的硫酸庆大霉素缓释微球

以生物可降解聚羟基丁酸酯和羟基戊酸酯的共聚物(PHBV)、乙交酯和丙交酯的无规共聚物(PLGA)两种高分子作为壁材,采用复乳溶剂挥发法,制备了包裹硫酸庆大霉素(GS)的载药微球。在扫描电镜下观察到所得微球表面呈多孔状,为球形或椭圆形,粒径在20~80μm。分析结果表明,包封率在60%以上,在体外16~20 d内药物全部释放,90 d微球体外降解50%左右。     

In vitro and in vivo characterization of huperzine a loaded microspheres made from end-group uncapped poly(d,l-lactide acid) and poly(d,l-lactide-co-glycolide acid)

The purpose of this work was to develop biodegradable microspheres for long term delivery of a potent acetyl cholinesterase inhibitor, huperzine A (Hup-A), which is of interest in the palliative treatment of Alzheimer's disease. Microspheres were successfully prepared with specifically end-group uncapped poly(d,l-lactide acid) and poly(d,l-lactide-co-glycolide acid) using a simple o/w solvent evaporation method. The morphology, particle size and size distribution, drug loading capacity, drug entrapment efficiency (EE) and in vitro drug release were studied in detail. It was found that the terminal group and the inherent viscosity (IV) of the polymers played key role in the drug encapsulation: higher EE was achieved with end-group uncapped and low IV polymers. In vitro drug release from microspheres made from the selected three kinds of polymers revealed sustained release of Hup-A without significant burst release. Preliminary pharmacokinetic study following subcutaneous injection of Hup-A loaded microspheres illustrated the sustained release of the drug over 6-8 weeks at clinically relevant doses in vivo. The studies demonstrated the feasibility of long term delivery of Hup-A using biodegradable microspheres.     

Effect of tris(hydroxymethyl) aminomethane on the phase behavior of poly(ethylene imine)/cinnamic acid conjugate and the release property of cubic phase containing the conjugate

The upper critical solution temperature (UCST) of poly(ethylene imine)/cinnamic acid (PEI/CA) conjugate decreased as the tris(hydroxymethyl) aminomethane (THMAM) concentration increased. On the optical micrographs of PEI/CA mixture solutions at 25°C, the microspheres were found when the THMAM concentration was 0?mM and 100?mM, but hardly found at 200?mM. Monoolein (MO) cubic phase containing PEI, CA, and THMAM exhibited a bilayer structure on the TEM micrograph. The release degree of methylene blue loaded in the cubic phase was not strongly dependent on THMAM concentration at 25°C and 37°C, but strongly dependent on the concentration at 50°C.     

Effects of lung surfactants on rifampicin release rate from monodisperse rifampicin-loaded PLGA microspheres

We have prepared inhalable and monodisperse poly(lactide-co-glycolide) (PLGA) microspheres targeting tubercle bacilli residing in alveolar macrophages. The effects of pulmonary surfactant on the rifampicin (RFP) release rate from RFP-loaded poly (lactide-co-glycolide) microspheres were studied. Also, those of their surface properties of RFP-loaded PLGA microspheres were studied. The RFP release from RFP/PLGA microspheres was accelerated by adsorption of pulmonary surfactant on the particle surface. The fastest RFP release rate was observed from pulmonary surfactant-adsorbed PLGA particles in pH 7.4 buffer solution compared with those in pH 4.0 buffer solution and saline solution. The slowest release rate was observed in the case when saline solution was used as dispersion phase of RFP/PLGA microspheres, although RFP release rate increased by the addition of pulmonary surfactant. From these results it is suggested that when RFP/PLGA microspheres are administrated by inhalation, the RFP release rates from the particles which are not taken up by alveolar macrophages and remain in the alveoli will be small. On the other hand, the RFP release rates and release amounts will be high after RFP/PLGA microspheres are taken up by alveolar macrophages existing in phagosomes, but they become relatively small after RFP/PLGA microspheres move into phagosome-lysosomes by the fusion of phagosomes with lysosomes. The absolute values of the electrophoretic mobility of PLGA microspheres increased by the adsorption of pulmonary surfactants on the surfaces of PLGA microspheres. By analyzing the experimental data using the soft-particle theory, it was indicated that the microspheres became 'softer' and the surface charge density of microspheres increases by the degradation. On the other hand, the surface of PLGA microspheres became harder and the electric charge density increased by the adsorption of pulmonary surfactant on the surfaces of PLGA microspheres. The changes in the surface charge density with degradation became larger by the adsorption of the lung surfactant on PLGA microsphere surfaces. It is considered that the changes in surface properties of PLGA microspheres affect their uptake efficiency by alveolar macrophage.     

A novel preparation method for microspheres by glycerol modified solid‐in‐oil‐in‐water multi‐emulsion

Biodegradable material poly(D, L ‐lactic‐co‐glycolic) acid (PLGA) plays an important role in drug‐sustained release systems. Here, we describe a glycerol modified solid‐in‐oil‐in‐water (m‐S/O/W) emulsion method for PLGA microspheres, in order to encapsulate proteins in PLGA by utilizing dextran glassy particles to protect the proteins from denaturing, unfolding, and aggregation during preparation and new external water phase to prevent the inner dextran glassy particles from leaking into the external water phase. External water phase containing 20, 40, 60, 80% glycerol showed that proteins released faster and more completely with increased glycerol content. According to their varied release profiles, microspheres of different formulations could be used to encapsulate vaccines or for delivering proteins over long‐term. Copyright © 2009 John Wiley & Sons, Ltd.     

Enhanced gentamicin loading and release of PLGA and PLHMGA microspheres by varying the formulation parameters

The purpose of this study was to develop a suitable formulation for gentamicin sulfate (GS) that gives a sustained release of the drug. Therefore this drug was loaded into poly(D,L-lactide-co-glycolide) (PLGA) and poly(lactic-co-hydroxymethyl glycolic acid) (PLHMGA) microspheres. The effects of various formulation parameters (ethanol, surfactant, osmotic value of the external phase, polymer type and concentration) on particle characteristics (size, loading and release) were investigated. The GS loaded microspheres were prepared using a double emulsion evaporation technique. The results demonstrate that neither ethanol nor surfactants had beneficial effects on the drug loading efficiency (around 4-10%). However, an increase in buffer concentration (and thus osmotic pressure) of the external phase resulted in a substantial increase of GS-loading (from 10 to 28%). Further, an increase of concentration of PLGA in DCM from 10% to 15/20% caused a 4-time increase of the drug loading. The best formulation identified in this study had a loading efficiency of around 70% resulting in PLGA microspheres with a 6% (w/w) loading. The particles showed a burst release of the drug depending on their porosity, followed by a phase of 35 days where hardly any release occurred. The drug was then slowly released for around 25 days likely due to degradation of the microspheres. The drug loading efficiency of GS in PLHMGA was not significantly different from PLGA microspheres (64%). The release of GS from PLHMGA microspheres was faster than that of PLGA because the degradation rate of PLHMGA is more rapid than PLGA. This study shows that prolonged release of gentamicin can be obtained by loading this drug into microspheres made of biodegradable aliphatic polyesters.     

生物可降解5-氟尿嘧啶载药微球的制备及性能研究

5-氟尿嘧啶(5-Fu)为水溶性嘧啶类抗代谢药,是治疗实体肿瘤的首选药物．但5-Fu毒性很大,血浆中停留半衰期t1／2仅为10～20min．为了减少氟尿嘧啶的毒副作用并提高药物利用率,可以将其制成聚合物载药微球．聚酯类高分子是较为常用的生物降解型药物载体材料,其中聚乳酸(PLA)及其共聚物具有良好的生物相容性及生物可降解性,常被广泛应用于药物缓释材料,     

A Polysaccharide-based Hydrogel and PLGA Microspheres for Sustained P24 Peptide Delivery: An In vitro and In vivo Study Based on Osteogenic Capability

A bone morphogenetic protein-2(BMP-2) derived synthetic oligopeptide, S [PO4]KIPKASSVPTELSAISTLYLDDD(P24), has shown great potential for facilitating bone regeneration. However, P24 cannot be directly used onto bone defects, while a continuous sustained delivery of P24 may lead to a better formation of bone tissue. Based on this issue, we have developed a sustained delivery system incorporating P24-loaded poly(lactide-co-glycolide)(PLGA) microspheres and nano-hydroxyapatite(n-HA) into the composite hydrogel. The P24-contained compound material was characterized with NMR, FTIR and SEM to demonstrate the fomiation of compound structure containing P24, PLGA and n-HA. A continuous drug release of P24 was observed for over 60 d that evidently enhanced the efficiency in promoting the proliferation of MC3T3-E1 cells and the secrete of alkaline phosphatase(ALP) in vitro. Moreover, the osteoinduction eflect of the hydrogel system with P24 peptide niicrospheres was demonstrated in vivo and manifested by the result of immunohistochemistry. This novel injectable composite hydrogel is expected to be applied to improving the bone defect treatment in bone tissue engineering.     

Preparation, characterization, and pharmacodynamics of exenatide-loaded poly(DL-lactic-co-glycolic acid) microspheres

Exenatide (synthetic exendin-4), a 39-amino acid peptide, was encapsulated in poly(DL-lactic-co-glycolic acid) (PLGA) microspheres as a sustained release delivery system for the therapy of type 2 diabetes mellitus. The microspheres were prepared by a double-emulsion solvent evaporation method and the particle size, surface morphology, drug encapsulation efficiency, in vitro release profiles and in vivo hypoglycemic activity were evaluated. The results indicated that the morphology of the exenatide PLGA microspheres presented as a spherical shape with smooth surface, and the particle sizes distributed from 5.8 to 13.6 μm. The drug encapsulation efficiency tested by micro-bicinchoninic acid (BCA) assay was influenced by certain parameters such as inner and outer aqueous phase volume, PLGA concentration in oil phase, polyvinyl alcohol (PVA) concentrations in outer aqueous phase. Moreover, in vitro release behaviors were also affected by some parameters such as polymer type, PLGA molecular, internal aqueous phase volume, PLGA concentration. The pharmacodynamics in streptozotocin (STZ)-induced diabetic mice suggested that, exenatide microspheres have a significant hypoglycemic activity within one month, and its controlling of plasma glucose was similar to that of exenatide solution injected twice daily with identical exenatide amount. In conclusion, this microsphere could be a well sustained delivery system for exenatide to treat type 2 diabetes mellitus.     

聚L-谷氨酸担载胰岛素口服微球的制备与评价

以聚L-谷氨酸为载体材料, 采用无水乳液法制备了口服胰岛素微球, 微球直径在5~20 μm, 载药质量分数为5%~9%. 载药微球具有良好的pH敏感释放行为, 在胃模拟液中2 h释放量约为5%, 在肠道模拟液中2 h释放90%以上. 考察聚合物分子量、溶液浓度、理论投药量及混合材料对微球释放行为的影响.     

A spheres-in-sphere structure for improving protein-loading poly (lactide-co-glycolide) microspheres

In present study, protein loaded poly (lactide-co-glycolide)/chitosan microspheres (PLGA/CS MSs) with spheres-in-sphere structure were prepared in order to weaken the burst release of protein from PLGA microspheres (PLGA MSs) and to buffer acidic micro-milieu. The PLGA MSs and PLGA/CS MSs were characterized in terms of their size distribution, morphology, drug-loading rate, zeta potential and physical-chemical properties. The incubation experiments of PLGA MSs and PLGA/CS MSs were manipulated in PBS solution at pH 7.4, 37 °C to monitor the release of BSA and the vehicles degradation. The release kinetic of BSA was illuminated mainly based on the degradation processes of the matrices. External CS crusts were proved to strikingly improve the release kinetic of the model protein by reducing initial burst release and extending continuous release while acting as a diffusion barrier. Moreover, using PLGA/CS MSs could avoid the decrease of pH value resulted from the acidic products of PLGA MSs because of the effective buffer action of the basic groups in CS. The results demonstrated that the spheres-in-sphere structure is an effective way to control the initial burst release of protein and to overcome the acidic problem of protein-loading PLGA MSs.     

Long-term controlled release of dual drugs from MBG/PLGA composite microspheres

In this study, a long-term controlled drug release system was designed based on mesoporous bioactive glass coated with poly(lactide-co-glycolide) (MBG/PLGA). In this system ibuprofen (Ibu) and egg white protein were used as the model drugs. Firstly, Ibu was loaded into MBG and MBG/PLGA microspheres were formed after MBG/PLGA. Then the egg white protein was adsorbed outside of the MBG/PLGA because of the interaction between the hydroxyapatite and the protein. The drug release tests indicate that Ibu and egg white protein can release from the long-term controlled dual drugs system at the same time. Notably, the release time of Ibu can reach 18 days, and the release time of egg white protein can reach to 6 days due to the role of PLGA. The release rate of Ibu is 49 % of loading rate (46 %), while the release rate of egg white protein is 47 % of adsorption value (184 μg/mg), indicating that the dual drug release system is highly potential in the practical bone repair application.     

Preparation and evaluation of nano-hydroxyapatite/poly(styrene-divinylbenzene) porous microsphere for aspirin carrier

A novel vehicle for the delivery of aspirin (ASA) was prepared from porous nano-hydroxyapatite/poly(styrene-divinylbenzene) [nano-HAP/P(St-DVB)] composite microspheres by grafting nano-HAP [Ca10(PO4)6(OH)2] onto porous P(St-DVB) microspheres. Four types of porous composite microspheres were prepared, each with different nano-HAP contents. The ASA-loaded composite microspheres prepared with 10% and 15% nano-HAP (mass ratio) exhibited excellent buoyancy with relatively short instantaneous floating time (within 10 min) and a long sustained floating time (12 h) in simulated gastric juice. They also offered good sustained release of ASA (up to 8 h). Furthermore, these composite microspheres displayed good buffering capacity that prevented the buildup of acidity caused by hydrolysis of ASA, keeping the pH of gastric juice within the normal range (pH 0.9 to 1.5). The results showed that porous nano-HAP/P(St-DVB) composite microspheres prepared with 10% and 15% nano-HAP could be used as a novel drug carrier for ASA, providing a sustained release dose without leading to stomach irritation, a side effect that is often associated with ASA medication.