Succinic semialdehyde reductase was inactivated by o-phthalaldehyde. The inactivation followed pseudo-first order kinetics, and the second-order rate constant for the inactivation process was 28 $M^{-1}s^{-1}$ at pH 7.4 and $25^{circ}C$. The absorption spectrum ($lambda_{max}$ 337 nm) and fluorescence excitation ($lambda_{max}$ 340 nm) and fluorescence emission spectra ($lambda_{max}$ 409 nm) were consistent with the formation of an isoindole derivative in the catalytic site between a cysteine and a lysine residue approximately about 3 $AA$ apart. The substrate, succinic semialdehyde, did not protect enzymatic activity against inactivation, whereas the coenzyme NADPH protected against o-phthaladehyde induced inactivation of the enzyme. About 1 isoindole group per mol of the enzyme was formed following complete loss of enzymatic activity. These results suggest that the amino acid residues of the enzyme participating in a reaction with o-phthalaldehyde are cysteinyl and lysyl residues at or near the NADPH binding site.