Study On The Cyclosporine A-loaded Eudragit L100 Nanoparticles

Objective: The cyclosporine A-loaded Eudragit L100 nanoparticles (CyA-L100-NP) were prepared by optimal technological procedure, its stability of storage, freeze-drying technique and the characteristic of drug release in vitro were investigated. The pharmacokinetics and distribution characteristics were also studied.Method: CyA-L100-NP were prepared by quasi-emulsion solvent diffusion technique,choosed Eudragit L100 as the carrier material and poloxamer 188 as the emulsifying agent . The uniform experimental design﹝U7(76)﹞ was applied to theoptimization of the formulation based on the particle size, the drug loading and entrapped efficiency etc. CyA-L100-NP was analyzed by dynamic light scattering for the particle size, transmission electron microscopy for morphology and x-ray diffractometer for physical characteristics. CyA-L100-NP were stored under 4,25,40℃during two months. The stability of the sample about the particle sizes , the content of CyA and the drug entrapped was investigated.The cryoprotectants of 2%, 4% and 6% trehalose, sucrose , lactose ,glucose, sorbitol and mannitol were studied in freeze-drying test in order to evaluating the protection of nanoparticles. The best cryoprotectant was chosen and the freeze-drying technique was established. CyA-L100-NP or Neoral were dispersed into the phosphate buffer containing 0.1% SDS at different pH values such as 1.0, 2.0, 3.0, 4.0, 5.0, 6.0, 6.8, 7.4 respectively. The defference of drug release between the two preparations was compared.120 mice ,♂, were oral administration Neoral or CyA-L100-NP at the dose of 25mg/kg. After oral administration, at the different times such as 0.25, 0.5, 1, 1.5, 2, 4, 6, 8, 12h, the blood was obtained by culling the eyeballs. After executing mice by cutting down their cervicums ,the heart ,liver, spleen, lung and kidney were rapidly removed ,grinded into homogenate, added internal standard Cyclosporine D(CyD), the drug was extracted by liquid-liquid extraction method and determined by HPLC. Their compartment models were fitting by 3p87 software and their pharmacokinetics parameters were calculated using trapezoidal rule. The targeting properties were evaluated.Results: The average particle size of nanoparticles was 39.75±0.50 nm, the yield was 92.21±2.06%, the drug entrapped efficiency was 99.57±0.04% and the drug loading was 15.37±0.34%. The nanoparticles had small size , were look around or ellipse and regular under TEM. The drug peak was disappeared in the x-ray diffraction pattern, showed that CyA was enveloped in the nanoparticles. The nanoparticle colloids solution could stored under 4,25℃during two months, without changing of the particle size, drug content and entrapped efficiency. The size of nanoparticles became bigger after 1 month under 40℃,showed that the solution can't be stored under high temperature.2% lactose was chosen as cryoprotectant, which had a good effecttion of freeze-drying. The average particle size of nanoparticles by freeze-thaw using water could be controlled at about 100nm. Neoral's releasing behavior had no significant difference at different pH value and the releasing rate was over 80% in 2h.The release rate of CyA-L100-NP was increased with the higher pH value .Two-compartment model(weight 1) was adaorive to describe the pharmacokinetic charateristics of Neoral and CyA-L100-NP in blood of mice. Compared with Neoral ,the relative bioavailability of CyA-L100-NP were 118.3%. The distribution experiment showed that CyA-L100-NP had no targeting effect compared to Neoral, but it changed the distribution of drug in vivo, the drug was dispersed more in spleen and lung, less in liver and kidney .It was supposed that it might be profit to reducing hepatotoxicity and nephrotoxicity of CyA.Conclusion: In this paper, the technique of preparing nanoparticles and freeze-drying was optimized systematically. The stable techonolgy of preparation was abtained. Based on the HPLC method and dependable sample treatment method of biological specimen, the pharmacokinetics and distribution characteristics of mice after oral administration were syntheticly assessed.This paper supported the theory foundation for the development and application of CyA-loaded nanoparticle drug delivery system.