Light-dependent NADPH:protochlorophyllide oxidoreductase catalyses the photoreduction of protochlorophyllide to chlorophyllide in chlorophyll biosynthetic pathway. In this NADPH-dependent reaction a proton and a hydride are transferred to C17-C18 double bond of protochlorophyllide. The intramolecular charge transfer state of protochlorophyllide is assumed to play an important role during photoreduction. This work could show that formation of this state accompanies a red shift of the C(13')=O carbonyl stretching mode of the isocyclic fifth ring of protochlorophyllide, indicating an increase in the carbonyl bond length which is possibly evoked by hyrogen-bonding interactions. Protein sites in the catalytic pocket handicap the formation of the triplet state of protochlorophyllide, which stands in direct competition with the formation of the intramolecular charge transfer state. Moreover, a strong blue shift in the excited-state absorption of the intramolecular charge transfer state was observed, indicating a state stabilization by the protein sites. This stabilization could be due to hydrogen-bonding interactions, too. Point mutations in the binding pocket of protochlorophyllide demonstrate a radical reduction of the enzymatic activity for the substitution of each amino acid. That is, the possiblility to form such hydrogen bonds is given for all amino acids in the binding pocket of protochlorophyllide. NADPH:protochlorophyllide oxidoreductase shows a higher efficiency after excitation with red light than with blue light. The result of this dependency on the excitation wavelength is a reduced population of the triplet state after excitation with red light.