Abstract: Compounds contain sulfur and their relevant derivatives are applied universally in the fields of chemistry, biochemistry, life science and so on. Among them, aryl thioethers are potential useful precursors for constructing various biological active of sulfur-containing heterocycles. Thus, researches for phenyl sulfides have received more and more concern. In this work, gas phase fragmentation reaction mechanism of three kinds of bis (phenylthio) benzene isomers was studied by atmospheric pressure chemical ionization in positive ion mode combined with ion trap mass spectrometer. Meanwhile, theoretical calculations were performed by the density functional theory (DFT) method at the B3LYP level with 6.31+G (d, p) basis set in the Gaussian 03 package to verify cleavage pathways. Under the positive ion mode, all compounds generated molecular ions firstly, and after collision-induced dissociation (CID), some interesting fragmentation behaviors were observed. Benzensulfur radical loss resulted from direct heterolytic cleavage of C—S bond and neutral elimination of HS radical were obtained among all compounds. For o-bis (phenylthio) benzenes, loss of HS radical was induced by radical delocalization, followed by cycloaddition and a serious of rearrangements. Moreover, continuous loss of HS radical appeared via similar fragmentation pathway with that of single HS radical loss. In addition, elimination of substituted benzenethiol was also detected, which was triggered by nucleophilic substitution, with H radical shift in the formed ion-neutral complex intermediate going after. For m-bis (phenylthio) benzenes, similar dissociation pattern leading to neutral elimination of HS radical was proposed, among which 1, 4-H proton transfer instead of thioketone-thiophenol interconversion resulted in substituted benzenethiol loss. In the case of neutral HS radical loss existed in p-bis (phenylthio) benzenes, reaction mechanism mediated by cycloaddition was illustrated as well. However, because of the linear chain structure, substituted benzenethiol could not be generated from p-bis (phenylthio) benzenes. Furthermore, distinctive elimination of S₂ triggered by phenyl transfer existed only in m and p-bis (phenylthio) benzenes due to smaller steric hindrance. There was also characteristic neutral loss of substituted benzene ring in m-bis (phenylthio) benzenes. As a result, o-bis (phenylthio) benzenes and the other two isomers could be differentiated according to the continuous HS radical and S₂ losses. While m and p-bis (phenylthio) benzenes could be differentiated based on the eliminations of substituted benzenethiol and substituted benzene. This work investigated reaction mechanism about main neutral loss of HS radical and substituted benzenethiol for three kinds of bis (phenylthio) benzene isomers in gas phase, using tandem mass spectrometry combined with theory calculation, which would provide support for further researches about sulfur-containing compounds. At the same time, discussion for the different fragmentation behaviors and characteristic neutral eliminations helps to realize differentiation and identification for bis(phenylthio) benzene isomers.