Synthesis and Characterization of PEGylated Liposomes and Nanostructured Lipid Carriers with Entrapped Bioactive Triterpenoids: Comparative Fingerprints and Quantification by UHPLC-QTOF-ESI+-MS, ATR-FTIR Spectroscopy, and HPLC-DAD

<b>Background/Objectives</b>: Pentacyclic triterpenoids, as bioactive phytochemicals, have proven to exhibit significant bioactivity (antioxidant, anti-inflammatory, hypoglycemic, and anticancer) and low cytotoxicity. This study developed convenient methods for extracting and characterizing a birch bark extract enriched in pentacyclic triterpenoids (betulin, betulinic acid, and lupeol) and entrapped in two bioavailable nanoformulations. The performance of ATR-FTIR spectroscopy as a cost-effective and non-destructive method was evaluated comparatively with accurate HPLC-based methods. <b>Methods</b>: The bark extract and pure betulin or betulinic acid were used to obtain PEGylated liposomes and nano lipid carriers (NLCs). Their size was characterized by light scattering diffraction. UV-Vis spectrometry was applied as a preliminary evaluation (1), as well as UHPLC-QTOF-ESI⁺-MS for structure identification (2), ATR-FTIR spectroscopy (for semi-quantitative evaluation) (3), and HPLC-DAD for an accurate quantification of each component, either in the organic solvents or in the nanoformulations (4). <b>Results</b>: The PEGylated liposomes had smaller sizes, and higher entrapment efficiency, significantly correlated between the three analytical methods. The performance of ATR-FTIR spectroscopy was positively correlated with HPLC-DAD data and confirmed the potential of this cheaper and reliable semi-quantitative method to evaluate the entrapment efficiency of TTs in liposome and NLC nanoformulations. <b>Conclusions</b>: The results recommend using liposomal nanoformulations for the entrapment of bioactive terpenoids and their characterization by ATR-FTIR after validation by HPLC-DAD. The ATR-FTIR spectroscopy also offers the possibility of screening in a short time different recipes of nanoformulations as well as their stability and bioavailability, which is useful for investigations in vitro and in vivo, which may confirm their efficacy as therapeutic agents.