Molecular Docking Study between Carmine Acid and β-Lactoglobulin: Interaction Mechanisms and Applications as a Bioactive Delivery System

Carmine acid (CA) is a natural red colorant extracted from female cochineal insects (Dactylopius coccus costa), widely used in the food industry due to its chemical stability. As a glycosylated anthraquinone derivative, CA shows resistance to pH variations, temperature, and light exposure, outperforming many synthetic and natural colorants. Its broad application in products such as yogurts, beverages, and processed meats is attributed to its ability to maintain vibrant colors even under harsh processing conditions. Among its main advantages are oxidative and thermal stability, approval by regulatory agencies such as the FDA, and high coloring strength at low concentrations. In this context, β-lactoglobulin (BLG), the main whey protein belonging to the lipocalin family, emerges as a promising carrier system. Its characteristic β-barrel structure contains a hydrophobic cavity ideal for binding small molecules, offering advantages such as biocompatibility, encapsulation capacity, and potential for controlled release. This study employed molecular docking techniques to investigate the interaction between CA and BLG under monomeric conditions, aiming to identify CA’s preferential binding site on BLG and assess the thermodynamic stability of the formed complex. The BLG structure (PDB ID: 3NPO, resolution 2.20 Å) was subjected to flexible docking using the MolDock algorithm in the Molegro Virtual Docker software, considering all 162 residues as flexible and including 54 crystallographic water molecules. Results revealed that 95% of the 600 evaluated conformations (20 runs × 30 poses) predominantly occupied the largest BLG cavity (79.872 ų), presenting significantly lower total energy (-20 kcal·mol⁻¹) compared to the smaller cavity (24.576 ų). The most stable conformation exhibited a total interaction energy of -130.658 kcal·mol⁻¹ and hydrogen bond energy of -18.4342 kcal·mol⁻¹. Eight hydrogen bond interactions were identified between CA and the BLG residues Asn88, Asn90, Asn109, Lys91, Glu108, and Ser116, with seven involving hydroxyl groups of CA. These findings confirm the thermodynamically favorable formation of the CA-BLG complex, supporting BLG's potential as a nanocarrier for CA. A detailed understanding of these molecular-level interactions opens perspectives for various technological applications, including the development of functional foods with protected natural colorants, controlled-release systems for pharmaceutical applications, and stable cosmetic formulations. Further molecular dynamics studies and experimental validation are needed to fully characterize the behavior of the complex under physiological and industrial processing conditions.