Amphotericin B Microemulsion Preparation And In Vitro Percutaneous Penetration Study

Objective In order to solve the drawback of poor solubility and infusion-related side effect for Amphotericin B (AmB), a microemulsion vehicle composed of isopropyl myristate (IPM), Tween80, isopropyl alcohol and distilled water for transdermal delivery was designed in this study. The purpose of this study was to explore a drug-loaded method with a powerful solubilization for AmB, and to find out the formula of microemulsion with the optimum skin permeability for AmB. The influence of pH value on content of drug-loaded, distribution of AmB in microemulsion and permeability of AmB microemulsion was investigated.Method During the preformulation study, the analytical method of high performance liquid chromatogram (HPLC) for AmB was established. The influential factor experiment of crude drug of AmB was carried out. The solubilities of AmB in distilled water and a series of phosphate buffers and the oil/water partition coefficients in n-octyl alcohol/water were determined, and the receptor mediums of percutaneous experiment for AmB were screened. During the preparation technology study, the types and proportions of surfactant, oil phase and cosurfactant were screened. The microstructure of microemulsions was identified by determining the conductivity and viscosity. Three different kind methods were tested to incorporate AmB into the microemulsion. The effects of structure of blank microemulsion and pH value on the drug quantity loaded in microemulsion were investigated. The solubilizing position of AmB in microemulsion and solubilizing mechanism of PIT (Phase Inversion Temperature) method were determined. The optimized formulation was characterized in terms of pH, viscosity, droplets dimensional distribution and mean droplet size. The stability of blank and drug loaded microemulsions was investigated by the centrifugalization test, high temperature test and accelerated test. During the vitro percutaneous permeation test, the effects of pH values and permeation enhancers on permeation rate were studied. Finally, the skin irritation test was carried out.Results The established HPLC method for AmB corresponded with the standards of analytics. The crude drug of AmB was easily influenced by the illumination. The solubility of AmB in water was 31.97μg/ml, which is a poor water soluble drug. The solubilities of AmB in buffer solutions depended on pH values. The oil/water partition coefficient in n-octyl alcohol/water was 56.95, which demonstrated that AmB possibly is an ideal model drug of cutaneous penetration. The receptor medium for AmB was selected as 20 % ethanol-pH7.4 buffer phosphate. Tween80, IPM and isopropyl alcohol were selected as surfactant, oil phase and cosurfactant respectively. At Km=1:1, O/SC<2:8 andФw with in the range of 55%~70%, the type of microemulsion determined by conductivity was O/W. The incorporation of AmB to the O/W microemulsion following the PIT method resulted in the maximum solubilization for AmB, and the content of AmB can attain 29.6 mg/10 ml. When the pH value was less than 5.2 or larger than 8.5, the drug quantity loaded in microemulsion had no obvious decrease. The strong solubilization for AmB based on PIT method can result in the increasing of hydrophilicity of AmB. The solubilized position of microemulsion for AmB was in the hydrophilic shell of the microemulsion droplets. Microemulsion A0 and A corresponded to the characteristics of microemulsion and were stable in the experiment conditions. During the vitro percutaneous permeation test, microemulsion M3 with appropriate pH (5.14) for skin showed the best percutaneous ability and was selected as optimized formula. The water-soluble azone was the good permeation enhancer for AmB and its anatonosis ratio was 5.167. The optimum percent of water-soluble azone in the optimized formula was 1%, which can increase the permeation rate of microemulsion M3 by 1.4 times. The optimal formula had no obvious visible stimulus on the normal skins of rabbits.Conclusion AmB possibly is an ideal model drug of cutaneous penetration. The incorporation of AmB to the O/W microemulsion following the PIT method resulted in the maximum solubilization for AmB. The adjustment of pH forced AmB converting from aqueous phase to the hydrophilic shell of the microemulsion droplets. At pH 5.14, optimized AmB microemulsion was stable in the condition of experiment, and exhibited the maxium permeation rate. The optimal formula for transdermal drug delivery of Amphotericin B was that Km-1:1, O/SC-1:9, aqueous phase-64%, water-soluble azone-1% and content of AmB-2.96 mg/ml.