Abstract: Two-center three-electron (2c3e) bonded disulfide complexes are of particular interest in biology and chemistry. This type of bonding is responsible for the stability of many ion-molecule clusters or long-lived radical intermediates widely encountered in many applications of organic chemistry, biology, and chemical catalysis. In this paper, disulfide-water radical cation complexes, derived from the gas-phase reaction between H₂O^+· and disulfides (RS-SR, R=methyl, ethyl, propyl, isopropyl or tert-butyl) were studied using tandem mass spectrometry and isotope-labelling experiment for exploring the characteristic of 2c3e bond inside these complexes．Interesting chemistry was revealed through the dissociation of RS-SR+H₂O^+· complexes. For example, CH₃S-SCH₃+H₂O^+· (m/z 112) would predominantly give product ions of m/z 66 and m/z 65 with 16% collision energy (CE), by the loss of CH₂=S (46 u) and CH₃S· (47 u), respectively, indicating the cleavage of the disulfide bond in the ionic complex of m/z 112. Similarly, when ethyl or propyl group replaced methyl group, RS-SR+H₂O^+· complexes would also produce the disulfide bond cleaved product. However, when isopropyl or tert-butyl group was in the disulfide, RS-SR+H₂O^+· would mainly generate RS-SR^+· by the loss of H₂O. It was proposed that RS-SR+H₂O^+· (R=methyl, ethyl, or propyl) with relative low steric hindrance mainly generated two ions of R₁SH+H₂O^+· and R₂SOH+H⁺, revealing that the bond strength between S and O was more stable than the bond strength between S and S in RS-SR+H₂O^+·. While RS-SR+H₂O^+· (R=isopropyl and tert-butyl) with large steric hindrance mostly produced ions of RS-SR^+· by loss of H₂O, suggesting that the bond strength between S and S was more stable than the bond strength between S and O in RS-SR+H₂O^+·. These results indicated that 2c3e bond in RS-SR+H₂O^+· with low steric hindrance should be the form of S∴S⁺, while 2c3e bond in RS-SR+H₂O+· with large steric hindrance should be the form of S∴O⁺. The experimental results may shed light on advanced studies about the 2c3e bonded disulfide-water radical complexes, better for understanding the life process involving 2c3e bonded disulfide in peptides and proteins.