Methodological advances in liposomal encapsulation efficiency determination: systematic review and analysis

Liposomes represent a highly promising drug delivery platform for a wide range of pharmaceutical compounds. Encapsulation efficiency (EE) stands as a critical quality attribute for liposomal formulations. Accurate determination of EE requires quantification of at least two parameters among the three distinct drug populations: total drug content, encapsulated drug fraction, and free drug concentration. However, due to the complex physicochemical characteristics of liposomes, particularly their structural flexibility, surface charge properties, and organic phase composition, direct measurement of encapsulated and free drug fractions presents significant analytical challenges. The ability to precisely quantify both free and total drug concentrations in liposomal formulations enables rapid and reliable evaluation of encapsulation efficiency, which is essential for guiding formulation optimisation and ensuring consistent product quality during scale-up manufacturing processes. This review provides a comprehensive analysis of various analytical techniques for EE determination, including (reverse) dialysis, ultrafiltration centrifugation, differential centrifugation (ultra/low-speed), and size exclusion chromatography, with particular emphasis on their methodological characteristics, applicable ranges, advantages, and limitations. Furthermore, we propose appropriate detection strategies for encapsulation efficiency assessment based on specific laboratory capabilities and the physicochemical properties of the investigational compounds.