This study investigated the in-season effect of intensified training comparing the efficacy of duration-matched intense intermittent exercise training with sprint interval training in increasing intermittent running performance, sprint ability, and muscle content of proteins related to ion handling and metabolism in football players. After the first two weeks in the season, 20 sub-elite football players completed either 10 weeks of intense intermittent training using the 10-20-30 training concept (10-20-30, n=12) or sprint interval training (SIT, n=10; work/rest-ratio: 6-s/54-s) three times weekly, with a ~20% reduction in weekly training time. Before and after the intervention, players performed a Yo-Yo intermittent recovery test level 1 (Yo-Yo IR1) and a 30-m sprint test. Furthermore, players had a muscle biopsy taken from the vastus lateralis. Yo-Yo IR1 performance increased by 330 m (95%CI: 178-482, P≤0.01) in 10-20-30, whereas no change was observed in SIT. Sprint time did not change in 10-20-30, but decreased by 0.04 s (95%CI: 0.00-0.09, P≤0.05) in SIT. Muscle content of HADHA (24%, P≤0.01), PDH-E1α (40%, P≤0.01), complex I-V of the electron transport chain (ETC) (51%, P≤0.01) and Na+ ,K+ -ATPase subunits α2 (33%, P≤0.05) and β1 (27%, P≤0.05) increased in 10-20-30, whereas content of DHPR (27%, P≤0.01) and complex I-V of the ETC (31%, P≤0.05) increased in SIT. Intense intermittent training, combining short sprints and a high aerobic load, is superior to regular sprint interval training in increasing intense intermittent running performance during a Yo-Yo IR1 test and muscle content of PDH-E1α and HADHA in sub-elite football players.