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Purpose: For the first time, we investigated the effects of altering cellular metabolic capacitance, via a 5-d creatine (Cr) loading protocol (20 g·d⁻¹), on oxygen uptake (VO2), accumulated oxygen deficit, muscle recruitment, and performance during a 1-km cycling time trial., Methods: In a double-blind, randomized, placebo-controlled design, 19 amateur cyclists were allocated to a Cr (n = 10, VO2peak = 56.0 ± 7.8 mL·kg⁻¹·min⁻¹) or placebo (n = 9, VO2peak = 56.0 ± 8.4 mL·kg⁻¹·min⁻¹) group, and performed a 1-km cycling time trial before and after the supplementation period., Results: Body mass was significantly increased in the Cr group (P < 0.05), but not in the placebo group. Participants adopted an "all-out" pacing strategy in both groups. However, Cr loading reduced VO2 immediately after the beginning (12th to 23th seconds), and this was accompanied by a reduced aerobic and increased anaerobic contribution. The VO2 mean response time was slower (pre: 17.2 ± 5.6 s vs post: 19.9 ± 4.6 s), the total O2 uptake was reduced (pre: 4.64 ± 0.59 L vs post: 4.47 ± 0.53 L), and the oxygen deficit was increased (pre: 0.82 ± 0.27 L vs post: 0.98 ± 0.25 L) after Cr loading. No differences were observed in the placebo group for these variables. Plasma lactate and integrated electromyography were not altered in either group, nor was the time to complete the trial (Cr group: pre: 89.1 ± 6.7 s vs post 89.1 ± 6.2 s and placebo group: pre 85.9 ± 4.9 s vs post 87.0 ± 5.4 s)., Conclusion: Cr loading slows the V˙O2 response and increases the anaerobic contribution during a 1-km cycling time trial.