A dehydrogenase is an enzyme belonging to the group of oxidoreductases that oxidizes a substrate by reducing an electron acceptor, usually NAD⁺/NADP⁺ or a flavin coenzyme such as FAD or FMN. Like all catalysts, they catalyze reverse as well as forward reactions, and in some cases this has physiological significance: for example, alcohol dehydrogenase catalyzes the oxidation of ethanol to acetaldehyde in animals, but in yeast it catalyzes the production of ethanol from acetaldehyde.

Oxidoreductases, enzymes that catalyze oxidation-reduction reactions, constitute Class EC 1 of the IUBMB classification of enzyme-catalyzed reactions. Any of these may be called dehydrogenases, especially those in which NAD⁺ is the electron acceptor (oxidant), but reductase is also used when the physiological emphasis on reduction of the substrate, and oxidase is used only when O₂ is the electron acceptor. The systematic name of an oxidoreductase is "donor:acceptor oxidoreductase", but, when possible, it is more conveniently named as "donor dehydrogenase".

Dehydrogenases oxidize a substrate by transferring hydrogen to an electron acceptor, common electron acceptors being NAD⁺ or FAD. This would be considered an oxidation of the substrate, in which the substrate either loses hydrogen atoms or gains an oxygen atom (from water). The name "dehydrogenase" is based on the idea that it facilitates the removal (de-) of hydrogen (-hydrogen-) and is an enzyme (-ase). Dehydrogenase reactions come most commonly in two forms: the transfer of a hydride and release of a proton (often with water as a second reactant), and the transfer of two hydrogens.

Sometimes a dehydrogenase catalyzed reaction will look like this: AH + B⁺ ↔ A⁺ + BH when a hydride is transferred.

A represents the substrate that will be oxidized, while B is the hydride acceptor. Note how when the hydride is transferred from A to B, the A has taken on a positive charge; this is because the enzyme has taken two electrons from the substrate in order to reduce the acceptor to BH.

The result of a dehydrogenase catalyzed reaction is not always the acquisition of a positive charge. Sometimes the substrate loses a proton. This may leave free electrons on the substrate that move into a double bond. This happens frequently when an alcohol is the substrate; when the proton on the oxygen leaves, the free electrons on the oxygen will be used to create a double bond, as seen in the oxidation of ethanol to acetaldehyde carried out by alcohol dehydrogenase in the image on the right.

Another possibility is that a water molecule will enter the reaction, contributing a hydroxide ion to the substrate and a proton to the environment. The net result on the substrate is the addition of one oxygen atom. This is seen for example in the oxidation of acetaldehyde to acetic acid by acetaldehyde dehydrogenase, a step in the metabolism of ethanol and in the production of vinegar.

In the above case, the dehydrogenase has transferred a hydride while releasing a proton, H⁺, but dehydrogenases can also transfer two hydrogens, using FAD as an electron acceptor. This would be depicted as AH₂ + B ↔ A + BH₂. A double bond is normally formed in between the two atoms that the hydrogens were taken from, as in the case of succinate dehydrogenase. The two hydrogens have been transferred to the carrier or the other product, with their electrons.