NAD(P)-dependent oxidoreductases are valuable tools for the synthesis of chiral compounds. Due to the high cost of the pyridine cofactors, in situ cofactor regeneration is required for preparative applications. In recent years, existing regeneration methodologies have been improved and new approaches have been devised. These include the use of newly discovered dehydrogenases that are stable in high contents of organic solvent and novel enzymes that can regenerate either the reduced or oxidized forms of the cofactor. The use of electrochemical methods has allowed cofactor regeneration for monooxygenases and natural or engineered whole-cell systems provide alternatives to approaches relying on purified enzymes.