Phosphorus–nitrogen compounds: Part 31. Syntheses, structural and stereogenic properties, in vitro cytotoxic and antimicrobial activities, and DNA interactions of bicyclotetraphosphazenes containing bulky side group
Graphical abstract
N4P4Cl8 was reacted with aminopodand to obtain 2,6-sas-phosphazene. The mono- and diamino-2,6-sas phosphazenes were prepared, and their characterizations were made. The interactions between the phosphazenes and pBR322 plasmid DNA, the evaluations for cytotoxic activity, and apoptotic and necrotic effects against A549 lung cancer and L929 fibroblast cell lines were studied.
Introduction
Phosphazene derivatives are hybrid molecules with an essentially linear and/or cyclic backbone of alternating phosphorous nitrogen atoms with the same and/or different organic side groups bonded to each phosphorous atom [1]. The chlorocyclophosphazenes, N3P3Cl6 and N4P4Cl8, are the best-known starting compounds and as such they have been extensively studied in the field of phosphazene chemistry [2], [3]. Although a large number of N4P4Cl8 derivatives have been synthesized with mono- and difunctional ligands [4], [5], [6], discussions of the substitution reaction patterns with polyfunctional ligands are very limited in the literature [7], [8], [9]. Hexachlorocyclotriphosphazene, N3P3Cl6, and octachlorocyclotetraphosphazene, N4P4Cl8, with bidentate and/or polydentate amines can produce spiro, ansa, dispiro (2,4- and 2,6-), trispiro, tetraspiro, spiro-ansa (2,4- and 2,6-), spiro-ansa-spiro (sas), bino, spiro-bino, and di(spiro-bino) products depending on the reaction conditions [10], [11], [12]. Up to now, two kinds of 2,6-bicyclo tetraphosphazene derivatives were obtained from the reactions of N4P4Cl8 with mono-functional amines (Fig. 1a) [13] and multi- functional reagents (Fig. 1b) [8].
A wide range of side groups (R1 and R2, Fig. 1a) may be bonded to this skeleton, leading to products with a similarly diverse range of physical and chemical properties [14]. Previously, to our knowledge there were only two papers about 2,4-sas and 2,6-sas bicyclophosphazenes in the literature [8], [9]. As part of our ongoing study of the reactions of N4P4Cl8 with multidendate ligands, we have concentrated primarily on the substituent exchange reactions of N4P4Cl8 with potassium {2,2′-[1,3-phenylenebis(methyleneiminomethylene)]diphenoxide, K2N2O2, (1a)} with the aim of obtaining bicyclotetraphosphazene derivatives and also exploring their biological activity. As known, mono- and polyamino substituted cyclotriphosphazene derivatives (e.g., aziridine, spermine and spermidine) have attracted a great deal of attention for their potential as anti-cancer agents [15], [16]. They exhibited cytotoxic activity against HT-29 (human colon adenocarcinoma), Hep2 (Human epidermoid carcinoma of the larynx), and Vero (African green monkey kidney) cells and stimulated apoptosis [17]. In addition, the antimicrobial activity of cyclotri- and tetraphosphazene derivatives was investigated against various bacteria and fungi [18], [9]. On the other hand, studies on the biological activity of cyclotetraphosphazene derivatives are very limited. It is known that octapyrrolidinocyclotetraphosphazene demonstrates significant anticancer activity [15]. The Cu(II) complex of a fully phenoxy-substituted star-branched cyclotetraphosphazene is active in the oxidative cleavage of DNA [19].
The present study focuses on the Cl replacement reactions of N4P4Cl8 with N2O2 tetradentate ligand (1) (Scheme 1) with the aim of obtaining the new 2,6-sas-bicyclotetraphosphazene derivatives, and to investigate the in vitro cytotoxic activity and apoptosis and necrosis effects against A549 lung cancer and L929 fibroblast cell lines. The evaluation of antimicrobial activity and DNA interactions of all the compounds were also presented.
Section snippets
Materials and method
All reactions were monitored using thin-layer chromatography (TLC) on Merck DC Alufolien Kiesegel 60 B254 sheets. Column chromatography was performed on Merck Kiesegel 60 (230–400 mesh ATSM) silica gel. The reactions were run out under argon atmosphere. Melting points were assessed with a Gallenkamp apparatus using a capillary tube. The Fourier transform infrared (FTIR) spectra were recorded on a Jasco FT/IR-430 spectrometer in KBr discs and reported in cm−1 units. One-dimensional (1D) 1H, 13C
Synthesis
The reaction of N3P3Cl6 with an equimolar amount of K2N2O2 (1a) in THF and toluene gave only sbs (2) product in a poor yield. When the reaction was carried out in acetonitrile, no product was isolated. However, the reaction of N4P4Cl8 with an equimolar amount of 1a in THF produces the partly (2,6-dispiro-bicyclo) substituted 2,6-sas compound (3) with 35% yield. The yield of 3 in acetonitrile was 3%, but the same compound was not obtained in toluene. The other expected products e.g., 2,4-sas, 2,4
Conclusions
The Cl replacement reactions of N4P4Cl8 with the dipotassium salt, K2N2O2 (1a) of ligand (1) gave 2,6-sas-cyclotetraphosphazene (3) in THF. When the reactions were made one equimolar amount of 3 with an excess mono- and diamines, along with the fully substituted 2,6-sas cyclotetraphosphazenes (4a, 4b and 5a–7a) were prepared. Although the excess morpholine and DASD were used in the reactions, the geminal bis- (4c and 4e) and tris- (4d and 4f) cyclotetraphosphazenes occurred. In addition, the
Acknowledgements
The authors thank the “Scientific and Technical Research Council of Turkey” (Grant No. 211T019), and Z.K. thanks Turkish Academy of Sciences (TÜBA) for partial support.
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