| Objects:To develop intravitreally injectable sustained-release microspheres for suppression of proliferative vitreoretinopathy(PVR). PVR is an intractable ophthalmopathy, which can cause retinal separation and even atrophia bulbi. The method applied is operation repair, but about 20% of the patients have the experienced resurgence of PVR. It was reported that intravitreal injection with the matrine solution could effectively suppress the inflammatory reaction and proliferation of fibroblasts. But the half-life of the drug is so short that frequent injection was needed to provide therapeutical concentration at the target site. A reasonable drug delivery formulation could be used to solve the problem. On the one hand, they can offer longer time period of drug concentration within therapeutic range than conventional formulations; On the other hand, they cause less side effects than the free drug solution. In this study, intravitreally injectable sustained-release microspheres were prepared, incorporating matrine as model drug and biodegradable chitosan as carrier, to improve bioavailability and decrease side effect, which could lay a foundation for its clinic use in the future. Methods: On the basis of scientific literatures and pretesting, chitosan microspheres were prepared using emulsion cross-linking technique with chitosan as carrier and glutaral as curing agent. We analyzed the formula factors and the process factors that had more influence to the evaluation indexes of microspheres. Then orthogonal design was used to select the optimal formula. In the course of that, the average diameter, the span, the drug-loading content and the entrapment efficiency of microspheres were selected as evaluation indexes, which were used to determine the best formula and process.The quality evaluation and vitro release of optimized microspheres were investigated. The surface morphology of microspheres was observed by scanning electron micrograph, and the diameter and size distribution was determined by optical microscope. In vitro release of matrine from microspheres was performed by oscillating in constant temperature method and the release data were analyzed with traditional models to study the release mechanism. The chemical and physical stability of optimal formula was investigated under following circumstances: high tempreture, high humidity, strong light, long natural store condition (25℃±2℃/RH60%±10% for 6 months).The pharmacokinetics and tissue distribution of matrine in ophthalmic tissues of rabbits were studied after intravitreal injection of Matrine-loaded microspheres and free Matrine solution. Ophthalmic tissues such as vitreous body, iris, retina and choroids, sclera were dissected at different times, using High-performance Liquid Chromatograph to detemine concentration of matrine in ophthalmic tissues. Cmax and Tmax were select from the true values. AUC0-∞, MRT and Frel were calculated using the trapezoid method.Rsults: The optimal formulation and process were defined through orthogonal experiments and the central composite design. It was found that ratio of the matrine and the chitosan,chitosan concentration and ratio of water to liquid paraffin had much more influence on indexes of microsphere. Microspheres made from the optimal formula have good sphericity and equal size distribution. The average diameter was 16.21±0.26μm,and adapted to intravitreally injectable. The evaluation indexes showed that the optimal formula and process had fine reproducity and stability. Drug-loading content, entrapment efficiency were 9.457±0.33% and 64.86±0.90%.The cumulative release from microspheres in vitro reached about 80.60±1.38% in 120 hours. The cumulative release profile could be distributed by Higuichi equation. The release mechanism showed that drug was released by small and complex holes in microspheres.Stability experiment: Under the condition of 60℃,RH75%±5%and strong light (4500±500lx) , the appearance, content and release characteristics of microspheres changed little. Under the condition of RH92.5%, the weight increasing ratios of microspheres were more than 5%. After storaged at 25℃±2℃/RH60%±10% for 6 months, microspheres had no significant difference in appearance, content and release characteristics.Tissue distribution and pharmacokinetics study: The microspheres showed lower concentrations but longer-term existence of the drug in all the tissues, and showed a remarkable bioavailability increase compared to free Matrine. In Matrine solution group, vitreous body got the peak concentration of Matrine 1 hour after injection, and could not determine at 12 hour. Other tissues were similar. In microspheres group, vitreous body got the peak concentration of Matrine at 24 hour, and could still determine at 120 hour. Other tissues were similar.Conclusions: The results showed that Matrine chitosan microspheres could prolong the residence time of drug in vitreous body and increase the bioavailability of Matrine, and moreover, this dosage form can be easily prepared and its quality can be control. Therefore it showes great potential in the ocular application.
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