CF₃PIP was synthesized through a modification of a previously reported procedure [29] -[31] . A mixture of 2- (Trifluoromethyl)benzaldehyde (0.69 mL, 5 m∙mol) and ammonium acetate (7.70 g, 0.1 mol) was added into a stirring solution of 1,10-Phenanthroline-5,6-dione (0.99 g, 5 m∙mol) in 60 mL of glacial acetic acid, and the mixture was continuously stirred at 60˚C for 6 hours. Then the deep red solution was cooled to room temperature and diluted with 100 mL distilled water. A grayish yellow precipitate was obtained by neutralization with ammonium hydroxide. Then the mixture was filtered and washed with water for three times. The crude solid power was purified by chromatography over 60 - 80 mesh SiO₂ using absolute ethanol as eluent. The solvent was removed and the products were collected, and dried in vacuo. m-CF₃PIP and p-CF₃PIP were prepared by a similar procedure with the compound CF₃PIP, with 3-(Trifluoromethyl)benzaldehyde (0.69 mL, 5 m∙mol) and 4-(Trifluoromethyl)benzaldehyde (0.69 ml, 5 m∙mol) in place of 2-(Trifluoromethyl)benzaldehyde (0.69 mL, 5 m∙mol) respectively, and a pink-yellow precipitate were obtained.

CF₃PIP: Yield: 61%. Anal. Found: C, 65.48; H, 3.14; N, 15.35; Calcd for C₂₀H₁₁F₃N₄: C, 65.93; H, 3.04; N, 15.28. ¹H NMR (DMSO-d₆, 500 MHz) δ: 9.03 (s, 2H, J = 8.7 Hz, ArH), 9.02 (s, 2H, J = 8.6 Hz, ArH), 8.47 (m, 1H, J = 8.3 Hz, ArH), 8.45 (m, 1H, J = 8.2 Hz, ArH), 7.96 (m, 1H, J = 8.1 Hz, ArH), 7.82 (br, 3H, J = 7.6 Hz, ArH), 3.32 (s, 1H, -NH). ¹³C NMR (DMSO-d₆, 500 MHz) δ: 148.9 C(15), 148.0 C(1,12), 143.7 C(5,14), 133.8 C(3ꞌ,5ꞌ), 129.7 C(1ꞌ), 129.5 C(3), 129.1 C(7), 126.6 C(2ꞌ,6ꞌ), 125.9 C(4ꞌ), 125.8 C(-CF₃), 124.8 C(8), 124.7 C(4), 123.4 C(9), 123.3 C(10), 122.6 C(6,13). ES-MS: (CH₃OH): m/z 365.0 ([M+H]⁺), 387.0 ([M+Na]⁺). IR (KBr disk): ν (cm⁻¹) = 3406.8 (s, N-H), 3078.2 (m, C-H), 1606.7 (s), 1551.8 (m), 1483.0 (s, C=C), 1315.3 (vs, C-F). Conductance (Ω⁻¹∙cm²∙mol⁻¹): 9.2.

m-CF₃PIP: Yield: 58%. Anal. Found: C, 65.67; H, 3.13; N, 15.42; Calcd for C₂₀H₁₁F₃N₄: C, 65.93; H, 3.04; N, 15.38. ¹H NMR (DMSO-d₆, 500 MHz) δ: 8.99 (m, 2H, J = 8.8 Hz, ArH), 8.83 (d, 2H, J = 8.7 Hz, ArH), 8.56 (m, 2H, J = 8.2 Hz, ArH), 7.78 (m, 1H, J = 8.0 Hz, ArH), 7.75 (m, 3H, J = 7.5 Hz, ArH). 3.38 (s, 1H, -NH). ¹³C NMR (DMSO-d₆, 500 MHz) δ: 148.2 C(15), 148.0 C(1,12), 143.6 C(5,14), 135.3 C(3ꞌ,5ꞌ), 132.5 C(1ꞌ), 130.3 C(3), 129.8 C(7), 128.2 C(2ꞌ,6ꞌ), 127.9 C(4ꞌ), 127.0 C(-CF3), 126.8 C(8), 126.7 C(4), 126.0 C(9), 124.8 C(10), 123.8 C(6,13). ES-MS: (CH₃OH): m/z 365.0 ([M+H]⁺), 387.0 ([M+Na]⁺). IR (KBr disk): ν (cm⁻¹) = 3241.6 (s, N-H), 3065.8, (m, C-H), 1607.2 (m), 1566.7 (s), 1548.5 (m, C=C), 1312.2 (vs, C-F). Conductance (Ω⁻¹∙ cm²∙mol⁻¹): 9.6.

p-CF₃PIP: Yield: 66%. Anal. Found: C, 65.78; H, 3.24; N, 15.32; Calcd for C₂₀H₁₁F₃N₄: C, 65.93; H, 3.04; N, 15.38. ¹H NMR (DMSO-d₆, 500 MHz) δ: 9.07 (d, 2H, J = 8.6 Hz, ArH), 8.84 (d, 2H, J = 8.4 Hz, ArH), 8.04 (d, 1H, J = 8.1 Hz, ArH), 7.97 (d, 1H, J = 7.8 Hz, ArH), 7.91 (d, 1H, J = 7.9 Hz, ArH), 7.83 (t, 3H, J = 7.7 Hz, ArH). 3.40 (s, 1H, -NH). ¹³C NMR (DMSO-d₆, 500 MHz) δ: 149.3 C(15), 147.7 C(1,12), 143.6 C(5,14), 131.6 C(3ꞌ,5ꞌ), 131.4 C(1ꞌ), 130.1 C(3), 129.9 C(7), 129.6 C(2ꞌ,6ꞌ), 128.4 C(4ꞌ), 125.5 C(-CF₃), 125.2 C(8), 123.2 C(4ꞌ), 123.1 C(4), 122.4 C(9), 122.3 C(10), 121.6 C(6,13). ES-MS: (CH₃OH): m/z 365.0 ([M+H]⁺), 387.0 ([M+Na]⁺). IR (KBr disk): ν (cm⁻¹) = 3371.2 (s, N-H), 3065.8 (w, C-H), 1608.2 (s), 1565.5 (m, C=C), 1326.5 (vs, C-F). Conductance (Ω⁻¹∙cm²∙mol⁻¹): 8.6.

[VO (Hbid) (CF₃PIP)] was prepared by the synthetic steps similar to a previously reported method [24] [32] [33] . A mixture of CF₃PIP (0.364 g, 1 m∙mol) and Hbid (0.266 g, 1 m∙mol) in absolute CH₃OH (60 mL) was heated at 80˚C under argon for 2 h. After dissolution, an aqueous solution (5 mL) of 1.0 m mol (0.217 g) quantity of VOSO₄∙xH2O was added dropwise to this mixture, and a breen precipitate was formed. The soild power was purified by recrystallization in methanol, and dried in vacuo. Yield: 36%. Anal. Found: C, 60.28; H, 2.92; N, 12.09; Calcd for C₃₅H₂₀F₃N₆O₄V: C, 60.35; H, 2.89; N, 12.07. ¹H-NMR (DMSO-d₆, 500 MHz) δ: 9.03 (s, 1H, CH=N), 8.68 (dd, 2H, ArH, J = 8.5 Hz), 8.60 (d, 2H, ArH, J = 8.4 Hz), 8.49 (br, 6H, ArH, J = 8.2 Hz), 8.26 (d, 1H, J = 8.4 Hz), 8.14 (d, 1H, ArH, J = 7.8 Hz), 7.82 (dd, 2H, ArH, J = 7.2 Hz), 7.14 (d, 2H, ArH, J = 6.8 Hz), 6.52 (dd, 2H, ArH, J = 7.8 Hz), 5.02 (s, 1H, -NH). ES-MS (CH₃OH): m/z: 695.1 ([M-H]⁻), 727.1 ([M+CH₃OH− H]⁻). IR (KBr disk): ν (cm⁻¹) = 3425.1 (N-H), 2973.3, 3077.0 (m, C=C-H), 1724.2 (vs, HC=N), 1608.0 (vs, C=O), 1314.1 (s, -CF₃), 974.1 (s, V=O), 730.8 (m, V-N). Conductance (Ω⁻¹∙cm²∙mol⁻¹): 10.7.

This compound was synthesized by a similar procedure as for the complex 1, with m-CF₃PIP (0.364 g, 1 m∙mol) in place of CF₃PIP. Yield: 43%. Anal. Found: C, 60.36; H, 2.91; N, 12.06; Calcd for C₃₅H₂₀F₃N₆O₄V: C, 60.35; H, 2.89; N, 12.07. ¹H NMR (DMSO-d₆, 500 MHz) δ: 8.87 (s, 1H, CH=N), 8.59 (dd, 2H, ArH, J = 8.5 Hz), 8.44 (d, 2H, ArH, J = 8.5 Hz), 8.36 (br, 6H, ArH, J = 8.3 Hz), 8.27 (d, 1H, J = 8.2 Hz), 8.08 (d, 1H, ArH, J = 7.7 Hz), 7.72 (dd, 2H, ArH, J = 7.8 Hz), 7.34 (d, 2H, ArH, J = 6.6 Hz), 6.48 (dd, 2H, ArH, J = 7.6 Hz), 5.20 (s, 1H, -NH). ES-MS (CH₃OH): m/z: 695.1 ([M-H]⁻), 727.1 ([M + CH₃OH − H]⁻). IR (KBr disk): ν (cm⁻¹)=3428.3 (N-H), 3027.0 (m, C=C-H), 1721.9 (s, HC=N), 1607.5 (vs, C=O), 1312.2 (s, -CF₃), 968.8 (s, V=O), 713.7 (m, V-N). Conductance (Ω⁻¹∙cm²∙mol⁻¹): 9.7.

This complex was prepared using a similar procedure described for complex 1. Yield: 36%. Anal. Found: C, 60.28; H, 2.93; N, 12.06; Calcd for C₃₅H₂₀F₃N₆O₄V: C, 60.35; H, 2.89; N, 12.07. ¹H-NMR (DMSO-d₆, 500MHz) δ: 9.12 (s, 1H, CH=N), 8.86 (d, 2H, ArH, J = 8.5 Hz), 8.63 (d, 2H, ArH, J = 8.4 Hz), 8.44 (br, 6H, ArH, J = 8.2 Hz), 8.26 (d, 1H, J = 8.4 Hz), 8.12 (d, 1H, ArH, J = 7.8 Hz), 7.80 (d, 2H, ArH, J = 7.2 Hz), 7.34 (d, 2H, ArH, J = 6.8 Hz), 6.62 (t, 2H, ArH, J = 7.8 Hz), 5.12 (s, 1H, -NH). ES-MS (CH₃OH): m/z: 695.1 ([M-H]⁻), 727.1 ([M + CH₃OH-H]⁻). IR (KBr disk): ν (cm⁻¹) = 3401.6 (N-H), 2982.9, 3077.2 (m, C=C-H), 1725.7 (vs, HC=N), 1609.4 (vs, C=O), 1314.5 (s, -CF₃), 956.6 (s, V=O), 730.9 (m, V-N). Conductance (Ω⁻¹∙cm²∙mol⁻¹): 11.8.

For the purpose of evaluating the capacity of oxovanadium complexes interacting with CT-DNA, the electronic spectra of complexes 1, 2 and 3 in the absence and presence of increasing amounts of CT-DNA are shown in Figure 1. As the CT-DNA concentration is increased, obvious hypochromism as well as a certain degree of bathochromism for the three oxovanadium complexes can be observed in the ultraviolet titration process at ca. 272 nm, which is attributed to π→π*. The hypochromism and bathochromism shift for 1 at 272 nm are 28.72% and 2 nm, respectively. In contrast, the hypochromism and bathochromism of complexes 2 and 3 are 20.93%, 1 nm and 30.36%, 2.5 nm, respectively. According to previously reported results [14] [23] -[25] , the UV-Vis spectral characteristics suggest that complexes 1-3 interact with CT-DNA most likely through a mode that involves a stacking interaction between the aromatic chromophore and the base pairs of DNA.

In order to elucidate the binding strengths of these complexes, the intrinsic binding constant K_b were calculated by monitoring the changes of absorbance in the ligand transfer bands with increasing amounts of CT-DNA. The K_b values of complexes 1-3 were calculated as 7.40 × 10⁴ (mol∙L⁻¹)⁻¹, 5.42 × 10⁴ (mol∙L⁻¹)⁻¹ and 1.25 × 10⁵ (mol∙L⁻¹)⁻¹, respectively, indicating that they strongly interact with DNA by intercalation. And the DNA-bind- ing activities of these three complexes are higher than that of reported mononuclear oxovanadium complexes VO(SAA)(phen) (4.50 × 10⁴ M⁻¹) and VO(MOSAA) (phen) (2.95 × 10⁴ M⁻¹) [25] , under our experimental conditions. The value of K_b increases in the order 2 < 1 < 3, this may due to the substituents of trifluoromethyl phenyl introduced on the phenanthroline ring of complexes 1-3, which may intercalate into base pairs of CT-DNA in complexes 1-3 will be altered, resulting in higher K_b. Meanwhile, the presence of an -CF₃ group on the aromatic moieties, which acts as an electron-drawing substituent on the intercalative ligand and thus gives stronger stacking interactions with DNA for complexes 1-3. And the electronic effects of substituted group at the different locations introduced on the aromatic ring of phenanthroline-based ligands may make differences in the DNA-binding affinities. The results also indicates an -CF₃ group substituted on the para-position of the aromatic exhibits stronger DNA-binding affinity than substituted on the ortho-position and meta-position.

The interaction of the complexes (20 μ∙mol∙L⁻¹) with CT-DNA was investigated using fluorescence emission titration experiment in the Tris buffer 1 at room temperature. The emission spectra of complexes 1, 2 and 3 in the absence and presence of CT-DNA exhibit luminescence in Tris buffer, with a maxima appearing at 442 nm, 443 nm and 440 nm respectively, which are shown in Figure 2. With increasing concentrations of CT-DNA, the emission intensities of complexes 1-3 grow to around by about 0.26, 0.19, and 0.36 times larger than those in the absence of CT-DNA and saturates at a [DNA]/[V] ratio of 20:1. The enhancement of emission intensity is indicative of binding of the complexes to the hydrophobic pocket of DNA, since the hydrophobic environment inside the DNA helix reduces the accessibility of water molecules to the complex and the complex mobility is restricted at the binding site, leading to decrease of the vibrational modes of relaxation [42] [43] . In addition, it also shows that 3 interacts with DNA more strongly than both 1 and 2, consistent with the UV-vis absorption titration spectra results.

To further clarify the nature of the binding interaction between both oxovanadium complexes and DNA in the absence of X-ray structural data or NMR spectra, viscosity measurements that are regarded as the least controversial and the most rigorous means of testing the binding model of DNA in solution [44] . It is well-known that a classical intercalation model leads to an apparent increase in viscosity of DNA solution due to base-pairs are pushed apart and hence an increase in overall DNA length [45] . In contrast, a partial, non-classical intercalation of compounds could bend (or kink) the DNA helix and reduce its effective length and, concomitantly, its viscosity [46] .

The effects of complexes 1, 2 and 3 on the viscosity of CT-DNA are shown in Figure 3. As can be seen in Figure 3, with amounts of the complexes increasing, the relative viscosity of DNA increases continuously to some extend. The results thus provide strong evidence for the interaction of complexes 1, 2 and 3 with CT-DNA by intercalation modes. Moreover, the increasing degree of viscosity is 2 < 1 < 3, which is consistent with electronic absorption spectra data, supporting the conclusion that complexes 1-3 interact with DNA by intercalation modes.

DNA melting experiments are generally applied to investigate the extent of intercalation, which were carried out by monitoring the intensity of DNA bases at 260 nm at different temperatures, both in the absence and presence of oxovanadium complexes. Thermal behaviors of DNA in the presence of compounds can give insight into their conformational changes when the temperature is raised and offer information about the interaction strength of complexes with DNA. As a rule, with the temperature in the solution rising, the double-stranded DNA will gradually dissociate to single strands and generate a hyperchromic effect on absorption spectra of DNA bases. Thus, the melting temperature T_m, which is defined as the temperature where half of the total base pairs are un- bounded, is usually introduced. Generally, T_m will increase considerable when intercalative binding occurs, since intercalation of the complexes into DNA base pairs causes stabilization of base stacking and hence raises the melting temperature of the double-stranded DNA [11] [24] [25] .

The melting curves of CT-DNA in the absence and presence of complexes 1-3 are shown in Figure 4; As shown in Figure 4, The T_m of CT-DNA in the absence of the complexes is 62.2˚C ± 0.2˚C, meanwhile, when the complexes at a concentration ratio [V]/[DNA] of 1:4, we can observe that T_m values in the presence of the complexes 1, 2 and 3 are 68.3˚C ± 0.2˚C, 65.3˚C ± 0.2˚C, 69.2˚C ± 0.2˚C, respectively. A comparison of the ΔT_m values of the complexes (6.1˚C, 3.1˚C and 7.0˚C for 1, 2, and 3, respectively) was agreement with those classical intercalators [47] [48] , which provided strong evidence for their binding with DNA by intercalation.

The experimental data reveals that complexes in collaboration with phenanthroline-substituted auxiliary li-

gands exhibit appreciable DNA intercalative activities, in addition, complex 3 showed a larger ΔT_m, indicating that 3 exhibits stronger DNA-binding affinity when interacting with DNA, in conformity with the electronic absorption titration.