Application And Study On Cyclosporine A Nano-Micelle

Objective:Currently, cyclosporine injection used in the clinic leads to a serious allergic reaction because of containing a large amount of Cremophor EL. Thus, this study focused on the preparation of a new type of cyclosporine A freeze-dried powder injection, studied on the composition and preparation technology, and investigated the storage stability of freeze-dried powder. Meanwhile, a serial of the physicochemical properties of the micelles were inspected, including the influence of accessories, drug loaded before and after, and freeze-drying process.Method:1. Detection of Cyclosporin A freeze-dried powder injection by UV spectrophotometryThis study employed UV spectrophotometer for detecting Cyclosporin A freeze-dried powder injection and the maximum absorption of Cyclosporin A, drawing the standard curve, carrying on the repetitive testing; Preparing a number of freeze-dried samples and measuring the Cyclosporin A according to the above method.2. Investigation of the composition of prescription screening and freeze-drying processThis part confirmed the best prescription cyclosporine micellar solution composition by determining dosages of Solutol HS15and ethanol, mixing time, bath temperature and other factors; Due to the prolonged storage of water injection will be cloudy, cohesion and so on, we freeze-dried them, and inspected the types and the use quantity of protective agents during the lyophilized drying process, thus drew the freeze-drying curve.3. Research of preparation technology and stability of freeze-dried samples. This part investigated the physicochemical properties of freeze-dried samples, including particle size and distribution, the effects of pH after lyophilization and reconstitution; Next, the stability of the samples was investigated, including dilution stability, pH stability, speed (40℃) and long term (25℃) stability and stress testing.4. Inspection of Micellization BehaviorThis part measured Solutol HS15critical micelle concentration (CMC) and aggregation number Nm by steady-state fluorescence probe method. Preparing blank micelles samples and inspecting the influence of ethanol towards CMC and Nm. Meanwhile, investigating the prescription dosage of cyclosporine A dosage and lyoprotectant different media dilution Solutol HS15critical micelle concentration (CMC).Results:1. The maximum absorption wavelength of cyclosporine A was206nm, and the formulation excipients had no absorption at this wavelength. The study established UV spectrophotometric method, which had good repeatability, high accuracy measurement results, showing that the concentration of cyclosporine A presented a good linear relationship from6μg/ml to200μg/ml, and the concentration of cyclosporine A was about18.65mg/mL.2. We initially employed the single factor method for determining the optimal preparation of drug-loaded micelles prescription including cyclosporine A250mg, ethanol650mg, Solutol HS153300mg, adding water volume to13ml. Then, we inspected alcohol sorbitol, glucose, lactose and sucrose, and ultimately decide Mannitol300mg conditions under the prescription,900mg sorbitol as a support during lyophilization agent.3. The average particle size of cyclosporin water reconstituted lyophilized sample was about22nm, pH was about6.18; Reconstitution time and the defoaming times were shorter (about2min). After four different dilution media were diluted to dissolve the lyophilized powder whose particle size measurements are about20nm, its acceleration and long-term tests showed good sample stability and could prepared in large-scale; The high temperature experiments and light has a little effect to the content and particle size, suggesting that they can storage under atmospheric temperature.4. The steady-state fluorescence probe determined the concentration of Solutol HS15critical micelle concentration (CMC) was0.217g/L. The amount of ethanol, cyclosporin A and lyophilized protective agent added to the prescription lead to the CMC of Solutol HS15increased, suggesting that these factors could effect the structure and properties of Solutol HS15micelle formation. In addition, after measuring the CMC of freeze-dried samples with different dilution media, we draw the conclusion that the order impacting Solutol HS15was0.9%NaCl>5%C6H12O6>10%C6H1206> H2O.