Modulation of thermomechanical-induced albumin aggregation by extracts of Centella asiatica

In this study, Centella asiatica extracts obtained through sequential extraction in ethyl acetate (CEE), methanol (CME), and water (CWE) were explored for the inhibition of thermal and thermomechanical-induced aggregation of albumin. Th-T fluorescence assay, dynamic light scattering (DLS), circular dichroism (CD) spectroscopy, and TEM analysis were used to study the aggregation behavior. Although CWE contained fewer compounds than CEE, it exhibits comparable inhibition characteristics as determined by High-Resolution Liquid Chromatography-Mass Spectrometry (HR-LCMS) and High-Performance Liquid Chromatography (HPLC) analyses. The rate of aggregation is decreased 1.64 to 0.96 h^-1 (i.e. two-fold) in the presence of extracts from under thermomechanical conditions. This effect correlates with the antioxidant capacity of the extract and is further supported by CD analysis, which showed no significant alteration in the native structure of BSA upon interaction with CWE, highlighting its aggregation-inhibiting potential. Isothermal titration calorimetry (ITC) identified the extract fraction with the highest binding affinity to BSA. Molecular docking studies revealed that chlorogenic acid, one of the most abundant compounds in CWE, interacts with albumin with the highest binding free energy (-9.0 kcal/mol), binding predominantly at Sudlow's sites I and III through hydrophobic interactions and hydrogen bonding. This study underscores the potential of natural compounds in preventing protein aggregation and provides a foundation for further investigation of lead bioactive components.