Porphyrin pyrrole sequencing: Low-energy collision-induced dissociation of (M + 7H) sup + generated in situ during ammonia chemical ionization

A method for pyrrole sequencing of porphyrins, based on high-pressure ammonia chemical ionization mass spectrometry/mass spectrometry (CI-MS/MS), is presented. High-pressure ammonia CI of porphyrins promotes formation of reduced molecular species and tri-, di-, and monopyrrolic fragment ions. The masses and pattern of the pyrrolic fragment ions can be used to determine the pyrrole sequence of the porphyrin. However, because of the complex ion source chemistry involved, these CI mass spectra are difficult to reproduce and are complicated by a plethora of structurally uninformative peaks. As such, pyrrole sequence elucidation based on the CI mass spectrum alone can be arduous, especially in the case of asymmetrically substituted porphyrins. In this paper, the MS/MS daughter ion spectrum of in situ formed (M + 7H){sup +} (i.e., the protonated porphyrinogen) is used to determine the porphyrin pyrrole sequence. Low-energy (30-100 eV) collision-induced dissociation (CID) of (M + 7H){sup +}, using a hybrid mass spectrometer of BEqQ geometry, yields, specific patterns of tri-, di-, and monopyrrolic daughter ions from which the pyrrole sequence of the porphyrin and combined mass of the substituents on the individual pyrroles can be determined. Several porphyrins of different pyrrole sequence are used to demonstrate the CI-MS/MS sequencing method. The capability of the CI-MS/MS method to sequence the pyrroles of porphyrins within a simple mixture of nonisobaric porphyrins is also demonstrated.