Breathing is not only the exchange of air in the lungs; at the cellular level it is a chain of oxidation–reduction reactions. In the 1920s Warburg used a self-built manometer, now called the Warburg apparatus, to measure oxygen consumption of tissue with high accuracy. He discovered that an iron-containing pigment protein, later called cytochrome, switches between oxidized and reduced forms to carry electrons and named it the “respiratory enzyme.” Warburg also proposed the presence of cytochrome oxidase that reacts with oxygen to form water at the end of the chain. These findings became the starting point for the idea of an electron transport chain. His data served as a standard when scientists discussed how ATP is produced. The work also influences medical studies of metabolic disorders such as the Warburg effect in cancer cells.

To quantify aerobic respiratory rates, Warburg modified constant-volume manometry, creating the temperature-controlled closed system now known as the Warburg apparatus. The device allowed real-time recording of oxygen consumption (QO2) in tissue extracts, while simultaneous absorption spectra revealed a reversible change near 570 nm that marked a heme protein. These proteins, later named cytochromes by David Keilin, possess stepwise redox potentials and pass electrons sequentially. Warburg proposed that cytochromes a and a3 catalyze the four-electron reduction of O2 to H2O and grouped them under the term “respiratory enzyme” (Atmungsferment). Stoichiometric equality between consumed oxygen and produced water supported the enzymatic nature of the reaction and foreshadowed the concept of an inner-mitochondrial electron transport chain. Adoption of the Warburg apparatus let scientists map sites of action for metabolic inhibitors and respiratory poisons, fostering progress in physiology and pharmacology. Observations of elevated glycolysis in tumor tissue later became known as the “Warburg effect,” a central topic in energy-metabolism research.

Warburg’s original papers analyzed difference spectra of cytochromes extracted from yeast and tumor tissues, identifying several ferro-ferri transitions with discrete midpoint potentials and correlating them with manometric QO2 data. He linked cyanide and carbon-monoxide inhibition of oxygen uptake to spectral changes in the α-band of cytochrome a3, assigning terminal oxidase activity to its heme–CuB center. Carbon-monoxide photodissociation provided on- and off-rate constants that became the first kinetic parameters for the oxidase reaction. The stoichiometry 2 H2 + O2 → 2 H2O was verified, ruling out peroxide intermediates under physiological conditions. Warburg postulated a vectorial proton translocation, anticipating Mitchell’s chemiosmotic hypothesis. His “respiration granules,” later recognized as mitochondria, retained redox activity after differential centrifugation. He introduced ADP-controlled respiration before Chance and Williams formalized state-3/state-4 transitions. The methodology enabled mapping of complexes I–IV and potentiometric titration of their redox potentials. Modern structural biology confirms that cytochrome c oxidase binds O2 at a binuclear heme-CuB center exactly as Warburg inferred from ligand inhibition patterns. His early metabolic control analysis foreshadowed systems-level studies and still provides benchmark data for pathogenic mitochondrial mutations.