Abstract: Cyclic peptides and their related species have been shown a unique class of molecules characterized by their biological function and structural diversity. Due to their biocompatibility and biostability in physiological environment, they have attracted the interest of many researchers in the fields of drug design and protein engineering. However, the structural characterization of cyclic peptides is much more challenging than that of linear peptides because of their complicated structures and stability. Mass spectrometry has great advantages of sensitivity and speed in identification of cyclic peptides, especially for complex mixture samples. Collisionally activated dissociation (CAD) mass spectrometry has contributed to linear peptide and protein sequencing, along with structural identification of post translational modifications. However, the structural analysis of cyclic peptides based on mass spectrometry is still a great challenge due to several difficulties, such as no termini apart from extended side chains, the existence of non-natural amino acids and the lack of corresponding database. Thus, understanding complicated fragmentation patterns of cyclic peptides is essential to characterize these compounds by mass spectrometry in different application scenarios. Mass spectrometry experiments are important for their structural analysis. In this work, sodiated and protonated ions of four cyclic peptide analogs were generated by electrospray ionization and were studied by tandem mass spectrometry. The CAD mass spectra of their sodiated and protonated ions showed a big difference. For the sodiated ions, the fragmentation pathway was characterized by the loss of the unit of CO in the first step, resulting the break of the ring and the followed sequential cleavages of the peptide. The sodium ion coordinated with backbone carbonyl oxygen atoms inside the ring, resulting their CAD mass spectra easy to be interpreted. The CAD mass spectra of protonated ions have several parallel dissociation channels, thus are more complicated. And this complexity is closely related to the functional groups on the cyclic peptide analogs. New fragmentation pathways were identified as the breaking of the bond of C-methyl ester. Compared with the protonation, the sodium complexation not only provided more easily interpretable spectra, but also improved sequence coverage. But different dissociation fragments from the protonated species might also provide some structural information that was valuable for unknown cyclic peptides and their related species. Density functional theory (DFT) calculations were applied for ACP1, revealing the different sites of protonation and metal complexation. It has been found that the undergoing informative dissociation in sodiated ions can mainly be attributed to the specific interaction between the coordinated sodium ion and backbone carbonyl oxygen atoms. The sodium complexation on cyclic peptides was helpful for understanding their MSn spectra and obtaining corresponding aminoacid sequences. The identification of unknown cyclic peptides in synthetic chemistry or natural product analysis can benefit from the combination of complementary product spectra generated from sodiated and protonated species.