Muscle contraction requires the physiology to adapt rapidly to meet the surge in energy demand. To investigate the shift in metabolic control, especially between oxygen and metabolism, researchers often depend on near infrared spectroscopy (NIRS) to measure non-invasively the tissue O₂. Because NIRS detects the overlapping myoglobin (Mb) and hemoglobin (Hb) signals in muscle, interpreting the data as an index of cellular or vascular O₂ requires deconvoluting the relative contribution. Currently, many in the NIRS field ascribe the signal to Hb. In contrast, ¹H NMR has only detected the Mb signal in contracting muscle, and comparative NIRS and NMR experiments indicate a predominant Mb contribution. The present study has examined the question of the NIRS signal origin by measuring simultaneously the ¹H, ³¹P NMR, and NIRS signals in finger flexor muscles during the transition from rest to contraction, recovery, ischemia, and reperfusion. The experiment results confirm a predominant Mb contribution to the NIRS signal from muscle. Given the NMR and NIRS corroborated changes in the intracellular O₂, the analysis shows that at the onset of muscle contraction, O₂ declines immediately and reaches new steady states as contraction intensity rises. Moreover, lactate formation increases even under quite aerobic condition.