Studies on pH gradient loading of amphoteric drug (Doxorubicin hydrochloride) into liposome

The present study was carried out with an aim to evaluate the factors influencing the active drug loading through transmembrane pH gradient using Ammonium Sulphate. (AS) The influence of various factors which influence the pH gradient loading of amphoteric drugs [Doxorubicin Hydrochloride (DH) in our case] such as incubation temperature, incubation time, transmembrane migration of ammonium ions, molarity which influence the pH gradient loading of amphoteric drug were investigated. Initially, optimization of AS (100mM) loading into HSPC liposomes by thin film hydration method were done and later influence of incubation time (0.5, 1, 2 and 3 hours) and incubation temperature (RT, 40, 50 and 60°C) on DH were investigated. Incubation time of 3 hours and incubation temperature of 60°C was found to be optimum for maximum loading of DH. AS of different molarity (20 to 300Mm) were loaded into HSPC and its effect on drug loading was investigated. Increase in molarity brought about the increase in loading of DH and liposome formulation AS (225Mm) gave maximum loading. Measurement of internal pH prior and after drug loading could correlate the influence of all the above parameters on the loading of DH through pH gradient method. Besides, physiochemical characterization of developed liposomes, in-vitro drug release studies, in-vivo pharmacokinetics studies of the prepared liposomal formulation and the related stability studies were also carried out.