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Objective To establish a detection method for the determination of tetrodotoxin (TTX) in sustained-release microspheres. Methods The HPLC separation of tetrodotoxin was performed on an Agilent ZORBAX SB-C18 column (4.6mm×150mm,5 μm) with acetonitrile, 8mmol/L sodium heptane sulfonate containing 0.005% TFA (5:95) (pH 4.0) as the mobile phase. The flow rate was 1.0 ml/min. The UV detection wavelength was 200 nm and the column temperature was 30 °C. Results The method had good specificity and linearity of TTX in the concentration range of 1−20 μg/ml. The intra-day precision, inter-day precision, stability and repeatability of the method were good, and the average recoveries were found between 98.0% and 102.0%. Conclusion This study established an HPLC method which was suitable for the determination of tetrodotoxin sustained-release microspheres. The method is accurate and reliable within the applicable range, with strong specificity, which could lead to quantitative detection.
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BACKGROUND: The repair of peripheral nerve defects by nerve conduit bridging can provide a suitable microenvironment for nerve regeneration. On one hand, it can provide a unique channel for nerve regeneration, prevent the invasion of peripheral connective tissue and the formation of scars. On the other hand, it can maintain endogenous and exogenous neurotrophic factors, growth factors and other stimulants to promote axon growth. OBJECTIVE: To observe the therapeutic effect of chitosan/polyvinyl alcohol catheter injected with brain-derived neurotrophic factor sustained-release microspheres to bridge peripheral nerve defects. METHODS: Chitosan/polyvinyl alcohol nerve conduit was prepared by repeated freeze-thaw technique. The brain-derived neurotrophic factor microspheres were obtained by polymer-alloys combined with oil-oil emulsion/solvent evaporation method. A 15 mm sciatic nerve defect model was made in the right hindlimb of 60 adult male Sprague-Dawley rats. They were selected and randomly divided into four groups (n=15 per group): group A implanted with autogenous sciatic nerve; group B implanted with chitosan/polyvinyl alcohol nerve catheter, injected with normal saline; group C implanted with chitosan/ polyvinyl alcohol nerve catheter, injected with brain-derived neurotrophic factor solution; group D implanted with chitosan/polyvinyl alcohol nerve catheter, injected with brain-derived neurotrophic factor sustained-release microspheres. General observation, histological inspection, and electrophysiological determination were performed at 4 months after the surgery. This study was approved by the Research Ethics Committee of the Second Hospital of Hebei Medical University. RESULTS AND CONCLUSION: (1) Gross anatomy showed that muscle atrophy in group A and group D was lighter than that in the other two groups. The grafts in four groups were all adhered to the peripheral tissues, and the nerve in the autotransplantation segment was strongly adhered to the peripheral tissues. In group D, the regenerated nerve had connected the distal and proximal nerves, and the regenerated nerve filled the conduit. (2) Electrophysiological examination showed that the latency of group D was shorter than that of groups B and C (P 0.05). (3) Histological observation showed that there were regenerated nerve fibers in groups B, C, and D. The diameter, number and thickness of myelin sheath of group D were larger than those of group B and group C (P 0.05). (4) The results showed that the injection of brain-derived neurotrophic factor microspheres into chitosan/PVA catheter had a long-term promoting effect on peripheral nerve regeneration.
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BACKGROUND: Transforming growth factor Β3/polylactic acid-glycolic acid (TGF-Β3/PLGA) sustained-release microspheres can maintain the effective drug concentration at the site of action and provide the feasibility for efficient utilization of growth factors. OBJECTIVE: To optimize the manufacturing process of TGF-Β3/PLGA sustained-release microspheres, and investigate their effects on the proliferation and migration of rabbit adipose-derived mesenchymal stem cells (ADSCs). METHODS: TGF-Β3/PLGA sustained-release microspheres were prepared by emulsification-solvent evaporation method. The morphology, particle size, drug spatial distribution, encapsulation efficiency, drug loading, and sustained release properties of the microspheres were characterized. The TGF-Β3/PLGA sustained-release microspheres were dissolved in phosphate buffered saline. The concentration of TGF-Β3 in the supernatant was detected at the corresponding time points. The microsphere morphology was observed by scanning electron microscopy at the corresponding time point. Adipose-derived mesenchymal stem cells were divided into six groups and then cultured with single culture medium (negative control) or culture medium containing TGF-Β3 or blank PLGA, or culture medium containing 10,100,1 000 g/L TGF-Β3/PLGA microspheres. Cell proliferation was detected by CCK-8 assay at the corresponding time point. Cells in each group were cultured for 24 hours with corresponding medium in a non-contact manner. The number of migratory cells was counted. RESULTS AND CONCLUSION: (1) TGF-Β3/PLGA sustained-release microspheres were spherical with smooth surface, no adhesion, and evenly distributed particle size. The microspheres had a diameter of 2-50 µm, and the protein drugs in the microspheres were evenly distributed, with high encapsulation efficacy and encapsulation dose. (2) The TGF-Β3/PLGA sustained-release microspheres had good degradation properties and were completely degraded after 6 months in vitro. At the same time, these microspheres had good sustained-release performance and released TGF-Β3 slowly for 45 days in vitro. (3) Blank microspheres and the sustained-release microspheres containing TGF-Β3 had no effect on the proliferation of adipose-derived mesenchymal stem cells. (4) Blank microspheres had no effect on the migration of adipose-derived mesenchymal stem cells, and the transforming growth factor 3 and the sustained-release microspheres containing TGF-Β3 promoted the migration of adipose-derived mesenchymal stem cells. There was no significant difference in the migration promotion between different concentrations of TGF-Β3. (5) These findings suggest that the TGF-Β3/PLGA sustained-release microspheres can promote the migration of adipose-derived mesenchymal stem cells without affecting their proliferation.
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OBJECTIVE: To obtain sustained-release indomethacin PLGA/Eudragit RS 100 microspheres for ophthalmic application and to evaluate its quality. METHODS: The microspheres were prepared by O/W solvent evaporation using PLGA and Eudragit RS 100. The morphological, drug content, entrapment efficiency, release behavior, particle size, residual organic solvent, Zeta potential of indomethacin PLGA/Eudragit RS 100 microspheres were determined; meanwhile, the cytotoxicity of blank microspheres was evaluated using PRE cells. RESULTS: Indomethacin microspheres containing PLGA and Eudragit RS 100 at the weight ratio of 1: 3 had even particle size distribution, and their surfaces were smooth under scan electronic microscope; and the mean diameters were (1676.4 ± 739.5) nm. The drug content and entrapment efficiency of the microspheres were (18.79 ± 0.19)% and (98.25 ± 2.11)%, respectively. Indomethacin was released slowly for a month from the microspheres, though burst release was observed within 24 h. The mean residual organic solvent was (267.33 ± 13.57) ppm. The zeta potential of all blank PLGA / Eudragit RS 100 microspheres was positive. The blank microspheres showed no sign of toxicity. CONCLUSION: Indomethacin PLGA / Eudragit RS 100 microspheres have a better clinical application prospect due to its high entrapment efficiency, narrow distribution, slow release and ocular safety. Copyright 2012 by the Chinese Pharmaceutical Association.
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OBJECTIVE:To optimize the preparation technology of Baicalein sustained release microspheres.METHODS:The microspheres containing baicalein were prepared using solvent evaporation technique.Multiple linear regression equation and quadratic polynomial equation were fitted respectively with the mass ratio of the matrix materials,sodium dodecyl sulfate(SDS)concentration in water phase and drug content as independent variables,and with the accumulative drug release percentage at 2,12 and 24 h(Y1,Y2,Y3)as dependent variables.RESULTS:The baicalein sustained release microspheres prepared in accordance with the above technology were spherical and even.The optimized technical conditions were as follows:the drug content was 25%;the ratio of the matrix materials was 3.5;SDS concentration was 0.08%;Y1,Y2 and Y3 were 21.05%,55.70% and 87.75%,respectively.All the three equations fitted using quadratic polynomial equations achieved good fitting results.CONCLUSION:Central composite design is convenient to use and it has satisfactory predictability thus deserving popularization.
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OBJECTIVE:To prepare triptolide-loaded sustained release microspheres and investigate its release profile in vivo and in vitro.METHODS:Triptolide-loaded sustained release microspheres were prepared by solvent evaporation method.A motionless method was used to investigate the process of release in vitro.The in vivo release process was investigated by pharmacokinetic experiment in rats.RESULTS:The microspheres were round and smooth examined by optical microscope.The size of triptolide loaded microspheres was(38.2?1.7) ?m and the encapsulation efficiency of triptolide loaded microspheres was(74.7?3.2)%.A constant in vitro release of triptolide-loaded microspheres was noted.The pharmacokinetic parameters of microspheres in vivo were as follows:Cmax was(114.7?31.90) ng?mL-1;tmax was(8.32?4.43) h;AUC was(1 774 282?1 046 152) ng?h?mL-1 and MRT was(596?165) h.In addition,the in vivo-in vitro correlation was good,r=0.955 3(P