Objective: The aim of the present study was to examine whether 10-20-30 training (consecutive 1-min intervals consisting of 30 s at low-speed, 20 s at moderate-speed, and 10 s at high-speed), performed with submaximal effort during the 10-s high-speed runs, would lead to improved performance as well as increased maximum oxygen uptake (VO₂-max) and muscle oxidative phosphorylation (OXPHOS). In addition, to examine to what extent the effects would compare to 10-20-30 running conducted with maximal effort. Design: Nineteen males were randomly assigned to 10-20-30 running performed with either submaximal (SUBMAX; n = 11) or maximal (MAX; n = 8) effort, which was conducted three times/week for 6 weeks (intervention; INT). Before and after INT, subjects completed a 5-km running test and a VO₂-max test, and a biopsy was obtained from m. vastus lateralis. Results: After compared to before INT, SUBMAX and MAX improved (p < 0.05) 5-km performance by 3.0% (20.8 ± 0.4 (means±SE) vs. 21.5 ± 0.4 min) and 2.3% (21.2 ± 0.4 vs. 21.6 ± 0.4 min), respectively, and VO₂-max was ~7% higher (p < 0.01) in both SUBMAX (57.0 ± 1.3 vs. 53.5 ± 1.1 mL/min/kg) and MAX (57.8 ± 1.2 vs. 53.7 ± 0.9 mL/min/kg), with no difference in the changes between groups. In SUBMAX, muscle OXPHOS was unchanged, whereas in MAX, muscle OXPHOS subunits (I-IV) and total OXPHOS (5.5 ± 0.3 vs 4.7 ± 0.3 A.U.) were 9%–29% higher (p < 0.05) after compared to before INT. Conclusion: Conducting 10-20-30 training with a non-maximal effort during the 10-s high-speed runs is as efficient in improving 5-km performance and VO₂-max as maximal effort exercise, whereas increase in muscle OXPHOS occur only when the 10-s high-speed runs are performed with maximal effort.