Phytoecdysteroids such as 20-hydroxyecdysone (20HE) are nutritional supplements marketed as enhancers of lean body mass. In this study the impact of 20HE ingestion on protein kinase B/Akt-mechanistic target of rapamycin complex 1 signaling in the skeletal muscle and liver of male rats was found to be limited. Bioavailability of 20HE, whether consumed alone or with leucine, also remained low at all doses ingested. Additional work is necessary to clarify 20HE mechanism of action in vivo.

Phytoecdysteroids such as 20HE are ingredients in various nutritional supplements marketed to enhance physical performance and lean body mass. Recent in vitro studies suggest 20HE may activate protein synthesis via a phosphoinositide‐3‐kinase‐Akt signaling mechanism. To confirm these findings in vivo and evaluate downstream activation of mammalian target of rapamycin complex 1 (mTORC1) signaling, overnight fasted 4‐6 week old male rats (n=5‐6 per group) were randomized to one of three study designs: 1) gavaged with 0, 10, 50, 200 mg/kg 20HE and euthanized 30 min post‐gavage; 2) gavaged with 200 mg/kg 20HE or excipient and euthanized at 30 min, 60 min, 120 min, 240 min post‐gavage, and 3) administered excipient or 200 mg/kg 20HE alone or in combination with 1.35 g/kg L‐leucine and euthanized 30 min post‐gavage. Independent evaluation of 20HE confirmed purity as reported by Sigma before solubilization in 3% DMSO or 70% Labrasol® as excipient. Phosphorylation of Akt at Thr308 and Ser473, mTOR at Ser2448, ribosomal S6 kinase (S6K1) at Thr389, and eukaryotic initiation factor 4E binding protein 1 (4E‐BP1) at Thr 37/46 were evaluated by immunoblot. Leucine administered in 3% DMSO or Labrasol robustly increased phosphorylation of mTOR, S6K1 and 4E‐BP1 but not Akt in muscle and liver. In contrast, at all doses and times, 20HE did not significantly increase phosphorylation of Akt, mTOR, S6K1 or 4E‐BP1 as compared to excipient controls. Furthermore, 20HE dampened mTORC1 signaling by leucine in both muscle and liver. These data do not support the idea that anabolic effects of 20HE are acutely mediated by mTORC1. Grant Funding Source: Supported by NJAES and USDA Multistate NC1184