Studies On Chitosan Modified Solid Lipid Nanoparticles Loaded With Methazolamide For The Ophthalmic Drug Delivery

ObjectiveThe aim of this study was to prepare methazolamide (MTZ) loaded solid lipidnanoparticles associated with biodegradable, non-toxic, water-soluble,low-molecular-weight and good biocompatible chitosan, for building a high stabilityand special targeting ocular drug delivery systems. In constrast, traditional solid lipidnanoparticles with negative charges are not conductive to corneal penetration ofdrugs.Methods1Water-soluble chitosan of low molecular weight were produced in the system ofacetic acid and hydrogen peroxide, using chitosans with5×104Da as materials. Theircorresponding molecular weights were further determined with Ubbelohde viscometer.Linear electrictitration method was adopted to detect the the degree of deacetylation(D.D) of these chitosans;2The chitosan modified solid lipid nanoparticles loaded with methazolamide (MTZ-CS-SLNs) were prepared according to a modified emulsion-solvent evaporationmethod. The optimal formulation was obtained by single factor evaluation,orthogonal design and Box-Behnken method, taking entrapment efficiency (EE%),drug loading (DL), particle diameter and Zeta potential as indicators. The stability ofMTZ-CS-SLNs was studied;3The physicochemical properties of MTZ-CS-SLNs were studied, such as pH,osmotic pressure, particle morphology, diameter, Zeta potential and so on. The matrixstate of MTZ-CS-SLNs was confirmed by Infrared spectroscopy (FI-IR), differential scanning calorimetry (DSC) and powder X-ray diffraction（XRD）. The drug releasebehavior was performed using the dialysis bag method. Besides, the cornealpermeability experiment was carried out using Franz diffusion pool;4The intraocular pressure (IOP)-lowering effects of different preparations wereevaluated using rabbits as animal model. The ocular tolerance and potential irritationof the preparations were evaluated according to a modified Draize test. Furthermore,the ocular toxicology were evaluated by histologic examination.Results1Water-soluble low molecular weight chitosan were successively prepared byhydrogen peroxide. The molecular weights degradated1.5h,2.5h,3.5h and4.5hwere10220Da,6569Da,4468Da and2251Da respectively. The degree ofdeacetylation (D.D) of chitosan were constant after degradation with hydrogenperoxide;2Optimal formulation of MTZ-CS-SLNs:20mg MTZ,100mg GMS,20mgphospholipids were dissolved in5mL ethanol at70℃to form oil phase. Theaqueous phase (1%Tween80containing1%PEG400, w/v) was heated to the sametemperature of the oil phase. The oil phase was instilled into the hot aqueous phaseunder mechanical stirring at1200rpm at70℃for dispersion. After removing theorganic solvent, the pre-emulsion was poured into the cold continuous phase (0-4℃)at a1:5v/v ratio under stirring at1000rpm and MTZ-CS-SLNs were obtained aftercontinual stirring for2h. The continuous phase was acetic acid solution whichcontained5%(w/v) mannitol and2.5mg/mL chitosan. The resulting suspensionswere filtered with0.45μm membrane. The preparation remained stable at4℃for4months;3The morphological examination of MTZ-CS-SLNs was regular sphere. Thenanoparticles have a particle diameter approximately250nm, PI0.291and a positive Zeta potential32.3mV. The average entrapment efficiency of the nanoparticles andtheir loading capacity were57.7%and9.8%respectively. Osmotic pressure and pHvalue were in compliance with the requirements of ocular drug formulations. FI-IR,DSC and XRD measurements suggested that the formation of MTZ-CS-SLNs. Thecumulative release percentage-time curve was fitted Weibull equation. The cornealpenetration ability of chitosan-modified nanoparticles is larger than that withoutchitosan modified nanoparticles;4In the experiments involving a single topical administration of MTZ-CS-SLNs inrabbits, IOP reduction could last over8h, which was significantly longer thanMTZ-SLNs and MTZ simulated artificial tear solution, similar with brinzolamide.Preparations used in this study showed no irritation, besides, the biopsy confirmed noocular toxicology in rabbit eyes.ConclusionThe preparation of MTZ-CS-SLNs was simple, which exhibited good stability,therapeutic efficacy, no-irritation and no occular toxicology. Compared withtraditional solid lipid nanoparticles, administration of MTZ-CS-SLNs could prolongthe duration of drug in the eye, improving drug permeation and bioavailability, whichhad good prospects for clinical application.