*4.1.1.5 Heating method*

Among all the conventional methods, the heating method is known to be the most attractive method for liposomes preparation due to its organic solvent free characteristics. In the heating method, lipids are hydrated for 1 hour, and heated for another hour above the transition temperature of the phospholipids in the presence of a hydrating agent (glycerin or propylene glycol 3%). When cholesterol is part of the formulation, the reaction medium is heated up to 100°C because of its high melting point. Being prepared under heating conditions, the resultant liposomes can be readily used without any further sterilization treatments, which minimizes both formulation complexity and timing. In addition, there is no need for further removal of the hydrating agents employed, since these represent physiologically acceptable ingredients that are well-established for pharmaceutical applications. Moreover, the observation that these hydrating agents can prevent particle coagulation and sedimentation makes them much more attractive as stabilizer and isotonizing additives. The hydroxyl groups of these hydrating agents provide a cryoprotective effect that makes the heating method an efficient method for the formulation of inhalable liposomes [41, 42].
