The condensation reactions of the tetrachloro mono (1 and 2) and bisferrocenylspirocyclotriphosphazenes (3-5) with 1,4-dioxa-8-azaspiro[4,5]decane (DASD) resulted in the formation of the partly and fully DASD-substituted phosphazenes. The reactions of equal amounts of 1-5 and DASD produced the mono-DASD-substituted ferrocenylphosphazenes (1a-5a), as the major product. When the reactions were carried out with 1 equiv of 1-5 and 2 equiv of DASD, corresponding geminal-phosphazenes (1b-5b) were isolated. Moreover, the reactions of 1 equiv of 1-5 and 3 equiv of DASD gave the tri-(1c-4c) and tetra-substituted (1d-5d) phosphazenes. When the excess DASD was used, the fully-substituted phosphazenes (1d-5d) were obtained. The chirality of 3a was evaluated using chiral HPLC column. The structures of all the phosphazenes were verified by FTIR, MS, H-1, C-13 and P-31 NMR, and HSQC spectral data. The crystal structures of 4a, 2b, 5b, and 1d were determined by X-ray diffraction techniques. The 10 phosphazene derivatives were screened for antimicrobial activity. Meanwhile, interactions between the compounds and pBR322 plasmid DNA were presented by agarose gel electrophoresis. The compounds 2b, 1d, 2d, and 4d were tested against HeLa cancer cell lines. Among these compounds, 4d had cytotoxic effect on HeLa cell after 24 h treatment. (C) 2013 Elsevier B. V. All rights reserved.