A tri-target in silico analysis of Testolift: a nutraceutical formulation targeting aromatase, myostatin, and prolyl hydroxylase-2 in testosterone regulation and muscle performance

Abstract

Abstract Declining testosterone levels and associated impairments in muscle function, endurance, and metabolic health represent a growing global concern, while limitations associated with long-term testosterone replacement therapy have intensified interest in safer, mechanism-based nutraceutical approaches. In the present study, a systems-oriented multi-target computational framework integrating physicochemical profiling, ADMET prediction, molecular docking, and molecular dynamics (MD) simulations was employed to investigate the mechanistic basis of the Testolift formulation composed of protodioscin, diosgenin, 5,7-dimethoxyflavone (DMF), and 5,7,4′-trimethoxyflavone (TMF). The phytochemicals were evaluated against three mechanistically interconnected targets namely aromatase (CYP19A1), myostatin, and prolyl hydroxylase-2 (PHD2), which are associated with testosterone regulation, muscle physiology, and oxygen-dependent metabolic adaptation. Protodioscin demonstrated extensive polar interactions and persistent hydrogen bonding within the CYP19A1 binding region, whereas diosgenin exhibited favorable lipophilicity and interaction characteristics consistent with steroidogenic signaling. DMF and TMF displayed the most favorable drug-like physicochemical profiles (QED = 0.74), high predicted intestinal absorption, and stable interactions with myostatin and PHD2. Molecular dynamics simulations performed over 150 ns confirmed stable ligand–protein complexes without major conformational destabilization. ADMET analysis further indicated complementary pharmacokinetic behavior together with an overall favorable predicted safety profile. Collectively, these findings support a coordinated multi-target nutraceutical framework in which structurally distinct phytochemicals modulate complementary peripheral pathways associated with testosterone balance, muscle anabolic regulation, and endurance-related metabolic adaptation, thereby supporting further experimental validation of the Testolift formulation.