Rosmarinic acid (RA), a polyphenolic compound abundantly found in the Lamiaceae family, has been shown to possess antioxidant and anti-inflammatory properties. In this study, we investigated whether it can promote energy expenditure in skeletal muscle cells. RA increased fatty acid oxidation and glucose utilization in C2C12 myotubes in a dose-dependent manner and increased the expression of Ca²⁺/calmodulin-dependent protein kinase kinase (CaMKK) and activated AMP-activated protein kinase (AMPK). CaMKK inhibitor attenuated the RA-induced AMPK activation and energy expenditure. Sirtuin1 (SIRT1) expression was enhanced by RA at both the gene and protein levels. We also observed that RA enhanced deacetylation of peroxisome proliferator-activated receptor γ coactivator-1α (PGC1α), accounting for the increase in energy expenditure-related genes. SIRT1 inhibitor attenuated the RA-induced PGC1α deacetylation and energy expenditure. Our findings suggests that CaMKK and SIRT1 play a role in the beneficial effects of RA on energy expenditure in skeletal muscle cells.