In the presence of oxygen, the charged pyruvate molecules — the final product of glycolysis, must enter the eukaryotic mitochondria to undergo pyruvate oxidation.

Since the enzymes for the reaction are present in the mitochondrial matrix, pyruvate needs to cross the mitochondrial double membrane.

Thus, it uses the porins present on the outer mitochondrial membrane and the mitochondrial pyruvate carrier on the inner membrane to reach the mitochondrial matrix.

Inside the matrix, it binds to the pyruvate dehydrogenase complex, composed of multiple copies of 3 types of enzymes.

In the first step of the reaction, the carboxyl group is removed from pyruvate and released as carbon dioxide.

The two-carbon molecule is oxidized to an acetyl group, and the released electrons reduce NAD⁺ into NADH and H⁺.

Finally, the acetyl group is transferred to coenzyme A, resulting in the formation of acetyl coenzyme A, which then enters the citric acid cycle to be further oxidized.