Combining lifestyle interventions such as intermittent fasting (IF) and high intensity interval training (HIIT) result in higher fat loss than IF and HIIT alone. However, the molecular pathways that underpin these changes are unclear. The present study investigated the effects of IF and/or HIIT on molecular markers of epididymal adipose tissue function. Eight-week-old male mice (C57BL/6) were initially fed a high fat (HF, 60%) and sugar (S, 30% w/v) diet for 12 weeks to induce obesity. They were then randomly allocated to four groups: IF (fasting for 2 alternate days/week), HIIT (3 days/week), combined IF+HIIT (2 alternate fasting days and 3 days HIIT) and CON (ad libitum HF/S diet) for 12 weeks. Body weight, fat mass (Echo MRI) and epididymal adipose tissue gene expression (qRT-PCR) were measured at the end of intervention period. At the end of intervention period, IF and IF+HIIT groups displayed significantly lower body weights (p<0.05), fat mass (p<0.05) and expression of the leptin gene (p<0.05) compared to both CON and HIIT groups. The IF+HIIT group displayed increased expression of genes related to fatty acid oxidation (Hydroxyacyl-coenzyme A dehydrogenase; (HADH), p<0.05) and intracellular trafficking (Fatty acid binding protein 4; FABP4), p<0.01)) compared to IF group only. A significant positive correlation (R=0.66, p<0.05) between HADH and FABP4 was also observed in IF+HIIT group. Simultaneous higher expression of both HADH and FABP4 suggests that combination of IF and HIIT has a synergistic effect on reduction in fat mass possibly due to increased fatty acid oxidation and mobilization. In summary, IF with or without HIIT are effective strategies for reducing fat mass accumulation potentially via different molecular signaling pathways within adipose tissue.