The Preparation And Application Of Montmorillonite-Betaxolol Hydrochloride Liposome For Ocular Delievry

In this research, an anti-glaucoma drugs, Betaxolol Hydrochloride(BH) was elected as model drug. The Mt-BH and Mt-BH loaded liposome(Mt-BH-LP) was prepared by acidification process and ethanol injection combined with ammonium sulfate gradient method, respectively. The preparation process, in vitro release, particle size and zeta potential, characterization, stability, safety, in vitro corneal permeability, precorneal residence ability and anti-glaucoma effect of Mt-BH-LP were researched.During the preparation process study, the optimal Mt-BH-LP was prepared by ethanol injection combined with ammonium sulfate gradient method with a feature of high encapsulation efficiency(75.85±2.15%) and drug-loading rate(11.41±0.29%). The in vitro release experiment shows that the BH solution released within 2.5h, while the regular BH liposome and Mt-BH-LP was only released 69.9% and 60.2% at 10 h, the Mt-BH-LP was more controlled-release. For microcosmic evaluation, the particle size and Zeta potential of regular BH liposome and Mt-BH-LP was 166.4±1.89 nm and19.33±0.41 m V, 218±22.32 nm and 17.03±0.25 m V, respectively. The addition of Mt-BH had a little effect on particle size and Zeta potential of Mt-BH-LP. The stability test indicated that the encapsulation efficiency and drug-loading rate of Mt-BH-LP was fell with the increase of time, and the low concentration of Ve had barely antioxidant effect on Mt-BH-LP, while a high concentration Ve caused sediment.According to the freeze-drying process study, 8% sucrose as cryoprotectant had a good protective effect. The entrapment rate, drug-loading rate and partical size of freeze-drying Mt-BH-LP was 70.17±1.13%, 10.12±0.68% and 247.6±4.22 nm, respectively. According to the morphology study, the regular BH liposome and Mt-BH-LP both smooth sphere that there had barely difference between them, indicating the addition of Mt-BH had little effect on the morphology of Mt-BH-LP. But the freeze-dried Mt-BH-LP had a rough surface, due to the effect of freeze-dry process. The particle size of regular BH liposome, Mt-BH-LP and freeze-dried Mt-BH-LP was about 180 nm, 220 nm and 210 nm respectively, which was similar to the results of Zeta potential and laser particle size analyzer.On the safety test, the chick embnyo corioallantoic membrane-trypan blue staining(CAM-TBS) test shows that the positive control group(Na OH) ariseed coagulation, and the weight of absorbed trypan blue was the greatest(about 8.8μg), revealing irritation of Na OH was the greatest. While the absorded BH of Mt-BH-LP was only about 0.71μg, having a little irritioan on CAM. The irritation of all samples was in this order: Na OH> BH solution> Betoptic> Mt-BH-LP> Normal Saline. The human immortalized cornea epithelial cell cytotoxicity test(MTT) shows that the Mt-BH-LP solution group had a high cell viability(>60%) under lower concentration, but the cell viability of Betoptic was all under 60% which might caused by its high viscidity and positive electricity. Hence, the cytotoxicity of Betoptic> BH solution> Mt-BH-LP> Blank liposome. According to the in vivo rabbit eye irritation, the Draize score of all samples all under 3, declaring the Mt-BH-LP had no observably irritation on eye. The cornea histologic section shows that the long-term administration of Mt-BH-LP had a little irritation on cornea. While all samples had different degree irritation on conjunctiva, due to its sensibility on exogenous compounds. But the irritation of BH solution and Betoptic were higher than Mt-BH-LP. According the results of 3 different irritation test, we could draw a conclusion that the safety of Mt-BH-LP was higher than BH solution and Betoptic, which had a potential application values on glaucoma therapy.In vitro corneal penetration experiments shows that the 6h cumulative diffusion amount(Q6h) of BH solution, Betoptic and Mt-BH-LP was 379.19, 54.21 and 28.89μg, respectively. The apparent permeability coefficient(Papp) of BH solution, Betoptic and Mt-BH-LP was 0.975×10-5, 0.13×10-5and 0.0726×10-5μg. cm.s-1, respectively. The change of formulation could slow the release of BH that could generate long-term anti-glaucoma effect and reduce the side effect. The release of BH from Mt-BH-LP including the release of free drug, drug in liposome, adsorbed drug on Mt and drug loaded into Mt. The characteristic release route(adsorbed drug on Mt and drug loaded into Mt) reduced the Papp and increased the slow release feature of Mt-BH-LP. The hydration level of Mt-BH-LP and Betoptic were in the normal range, while the BH solution was over 83%, revealing that the irritation of Mt-BH-LP was lower than BH solution. This result was consistent with safety test, the irritation of Mt-BH-LP was lower than BH solution and Betoptic.The tears elimination study shows that the precorneal residence time of BH solution, Mt-BH-LP and Betoptic was 60, 240 and 300 min, respectively. The BH concentration of Betoptic group declined slowly due to its high viscosity and retention ability. The precorneal residence ability Betoptic> Mt-BH-LP> BH solution. And there was the some order in anti-glaucoma study, the minimum IOP of BH solution, Mt-BH-LP and Betoptic was occurred at 60, 120 and 240 min, respectively. The IOP sharply regained to normal high IOP in the BH solution group after achieved the minimum IOP, while the anti-glaucoma effect of Mt-BH-LP and Betoptic chould last over 6 hours. The anti-glaucoma ability Betoptic> Mt-BH-LP> BH solution, which was consistent with the result of precorneal residence ability. The precorneal residence ability was conducive to the anti-glaucoma effect, decreasing administration frequency, toxic and side effect.