Integrated Multilevel Experimental Assessment of Heart Support Oil Phytocompounds Across Molecular Motor Proteins, β2-Adrenergic Signalling, Mitochondrial Stress, and Cardiomyocyte Contractile Function

Heart Support Oil (HSO) is a multicomponent botanical formulation derived from Juglans regia, Lycium barbarum, Citrus × paradisi, and Origanum vulgare. This study addressed two related but analytically distinct questions: whether selected analytically detectable HSO phytocompounds show direct target-engagement signals at molecular motor proteins and the β2-adrenergic receptor (β2-AR), and whether the complete standardised formulation modifies oxidative- and adrenergic-stress phenotypes in cardiomyocyte models. Selected constituents were examined by target-specific biophysical/biochemical assays, whereas complete HSO was evaluated in human induced-pluripotent-stem-cell-derived cardiomyocytes and AC16 cells using membrane-injury, oxidative-stress, mitochondrial, calcium-transient, electrophysiological, contractile, qRT-PCR, and immunoblot endpoints. HSO produced concentration-dependent improvements under the tested stress conditions. At 20 µg/mL, CellTiter-Glo signal reached 91.0 ± 1.4% of control, reactive oxygen species decreased to 1.29 ± 0.04-fold, and mitochondrial membrane potential and cellular ATP recovered to 89.8 ± 1.6% and 87.3 ± 1.3% of control, respectively. HSO also moderated isoproterenol-associated abnormalities in cAMP signalling, beat rate, calcium handling, and contractility and shifted stress-, inflammatory-, apoptotic-, and antioxidant-response markers toward control values. Direct target assays demonstrated measurable binding and enzymatic changes for selected constituents, but these molecular observations were not interpreted as proving that the same targets caused the formulation-level cardiomyocyte phenotype. The findings support further independently replicated, pharmacokinetic, mechanistic loss-of-function, and in vivo studies; they do not establish a unique causal mechanism or clinical efficacy.