Design, development and characterization of Papain-loaded nanostructured lipid carriers for enhanced stability and bio-accessibility in acidic environments

Papain, as potential therapeutic regime, can be utilized for managing specific diseased conditions. However, its application has been limited due to its natural instability; especially when exposed to an acidic environment with a pH lower than 2.8. Therefore, the purpose of this study is to develop a Papain-loaded Nanostructured Lipid Carriers (Papain-NLC) using the Box-Behnken design (BBD) to explore the influence of different variables on the response. In this study, an optimized model was formulated based on predicted responses and the actual results closely aligned with the predicted values. Scanning Electron Microscope (SEM) and Fourier -transform infrared spectroscopy (FT-IR) were used to analyze particle size, shape, entrapment efficiency, and drug release in order to learn more about the interactions between the formulation's components. In vitro release studies were performed, revealing a slow release of papain from the lipid matrix, coupled with the gradual degradation of matrix. Mathematical release kinetics, specifically the first -order kinetics model effectively described this release behaviour. In vitro bioaccessibility of Papain-NLCs using static digestion model was done and samples were evaluated by LC-MS/MS analysis. The results demonstrated that the static digestion model improved the bioaccessibility of Papain-NLCs. To ensure the chemical stability of Papain-NLCs, a six-month stability study was conducted, assessing particle size, polydispersity index, zeta potential and entrapment efficiency. Remarkably, no significant changes were observed, indicating the formulations robustness and potential for practical applications. Safety evaluation of the Papain-NLC's were performed using CaCo2 cell lines, employing MTT assay for cell viability and LDH assay for cell toxicity assessment.