The aim of this study was to assess the correlation between lower limb muscle activation, co-activation characteristics, and running economy (RE). Twenty-nine male recreational runners participated in two sessions: during the first session, body composition and RE at 10 km·h−1 were measured, while during the second session, we assessed muscle activation using sEMG for rectus femoris (RF), vastus lateralis (VL), vastus medialis (VM), biceps femoris (BF), gluteus maximus (GM), tibialis anterior (TA), gastrocnemius lateralis (LG), gastrocnemius medialis (MG), and soleus (SOL) muscles. Additionally, seven pairs of agonist-antagonist co-activation ratios were calculated. Pearson's and statistical non-parameters mapping (SnPM) correlation analyses were used to evaluate the muscular activity and RE, respectively. Pearson’s analysis showed that GM, VM, VL, LG, MG, and SOL activation during the stance phase was negatively correlated with RE (r = −0.499 to −0.592, p < 0.05), whereas RF and VL activation during the initial swing phase was positively correlated with RE (r = 0.522, r = 0.527; p < 0.05). Regarding muscle co-activation, the SOL-TA in the propulsion, the BF-VL, and GM-RF during the initial swing phase were negatively correlated with RE (r = −0.506 to −0.634, p < 0.05). In contrast, the RF-BF in the terminal swing phase was positively correlated with RE (r = 0.578, p < 0.05). Unfortunately, SnPM analysis did not show significant results. In conclusion, greater lower limb muscle activation during the stance phase is associated with better RE. During the swing phase, the co-activation of the high activation of the posterior thigh muscles (GM, BF) and the lower activation of the anterior thigh muscles (quadriceps) is associated with better RE.