Third generation fluoroquinolones antibacterial drug based mixed-ligand Cu(II) complexes: structure, antibacterial activity, superoxide dismutase activity and DNA–interaction approach

Figures & data

Table 1.  Physicochemical parameters and elemental details of the complexes.

Empirical formula for complexes	Elemental analysis% required (found)				Mp °C	% Yield	µeff·BM	λmaxnm	Formula weight (g/mol)
	C	H	N	Cu
C29H40ClCuF2N7O8 (1)	46.34(46.23)	5.36(5.85)	13.04(12.95)	8.45(8.39)	>300	75.44	1.77	668	750.19
C24H35ClCuF2N4O9S (2)	41.62(41.58)	5.09(5.01)	8.09(8.16)	9.17(9.19)	294	87.66	1.89	662	691.11
C25H36ClCuF2N5O9 (3)	43.67(43.74)	5.28(5.19)	10.19(10.02)	9.24(9.67)	>300	87.53	1.81	671	686.15
C27H38ClCuFN6O8 (4)	46.82(46.83)	5.53(5.54)	12.13(12.12)	9.17(9.16)	298	78.32	1.74	659	691.17
C22H34ClCuFN3O9S (5)	41.64(41.59)	5.4(5.38)	6.62(6.66)	10.01(9.97)	276	81.04	1.83	663	633.1
C23H34ClCuFN4O9 (6)	43.95(43.92)	5.45(5.42)	8.91(8.93)	10.11(10.12)	292	77.97	1.79	657	627.13

Figure 1.  Structure of the title complex [Cu(SPF)(A¹)Cl].5H₂O [1].

Table 2.  Spectrophotometric titration data for the complexes.

Volume of 0.001 M EDTA (mL)	Absorption at 745 nm
	(1)	(2)	(3)	(4)	(5)	(6)
0.35	0.007	0.006	0.004	0.01	0.006	0.008
0.7	0.012	0.009	0.007	0.014	0.011	0.012
1.05	0.016	0.014	0.011	0.018	0.016	0.016
1.4	0.02	0.017	0.014	0.021	0.02	0.02
1.75	0.025	0.02	0.018	0.024	0.026	0.024
2.1	0.029	0.024	0.021	0.027	0.029	0.028
2.45	0.033	0.028	0.025	0.032	0.034	0.032
2.8	0.035	0.031	0.028	0.035	0.036	0.035
3.15	0.036	0.033	0.03	0.036	0.036	0.036
3.5	0.036	0.033	0.03	0.036	0.036	0.036
Amount of complex dissolved in 20 ml (g)	0.02012	0.02064	0.02008	0.01989	0.01642	0.01792
Equivalents volume of 0.001 M EDTA (mL)	2.667	2.98556	2.9234	2.866	2.594	2.844
Equivalent Concentration of EDTA at end point (M)	0.0002667	0.000298556	0.00029234	0.0002866	0.0002594	0.0002844
Amount of Cu in 2 mL (g)	0.000169477	0.00018972	0.00018577	0.000182123	0.00016484	0.000180725
Amount of Cu in 20 mL (g)	0.001694772	0.001897204	0.0018577	0.001821228	0.00164838	0.001807248
% Cu experimental	8.42	9.19	9.25	9.16	10.04	10.09
% Cu theoretically	8.45	9.17	9.24	9.17	10.01	10.11

Figure 2.  Experimental cure from the spectrophotometric determination of metal content of 20.12 mg complex [Cu(SPF)(A¹)Cl].5H₂O dissolved in 20 ml of DD water by EDTA.

Table 3.  Characteristic absorptions bands of IR spectra of the complexes and drugs (cm⁻¹).

Comp	ν(C=O)pyridone	ν(H–O)Carboxyl	ν(COO)asy	ν(COO)sym	Δν	ν(M–N)	ν(M–O)	ν(M–S)
SPF	1728	3526	1620	1332	288	–	–	–
PFLH	1716	3518	1632	1308	324
1	1623	–	1585	1346	236	536	518	–
2	1625	–	1581	1351	225	534	514	432
3	1621	–	1579	1342	232	546	515	–
4	1627	–	1583	1344	239	539	521
5	1624	–	1589	1355	234	537	512	436
6	1617	–	1577	1342	235	544	517

Figure 3.  The FAB-mass spectrum of [Cu(SPF)(A¹)Cl].5H₂O at an accelerating voltage of 10 kV.

Figure 4.  Proposed fragmentation pattern for [Cu(SPF)(A¹)Cl].5H₂O complex.

Table 4.  MIC in terms of µM.

Compounds	Gram positive		Gram negative
	S. aureus	B. subtilis	S. marcescens	P. aeruginosa	E. coli
CuCl2·2H2O	2698	2815	2756	2404	3402
Sparfloxacin(SPF)	1.3	2.0	1.5	1.5	1.3
Pefloxacin(PFLH)	2.1	2.4	5.1	5.7	2.7
A^1	3212	3271	3212	3183	3154
A^2	>10000	>10000	>10000	>10000	>10000
A^3	3821	3414	3333	2902	3739
[Cu(SPF)(A^1)Cl].5H2O (1)	1.8	1.8	5.5	2.7	2.7
[Cu(SPF)(A^2)Cl].5H2O (2)	1.7	1.7	10.5	6.2	6.1
[Cu(SPF)(A^3)Cl].5H2O (3)	0.9	1.2	4.4	3.5	13.9
[Cu(PFL)(A^1)Cl].5H2O (4)	0.9	1.2	0.9	0.3	0.3
[Cu(PFL)(A^2)Cl].5H2O (5)	1.6	1.6	1.9	1.7	1.7
[Cu(PFL)(A^3)Cl].5H2O (6)	0.9	3.5	2.7	0.9	3.5

Table 5.  The binding constants (K_b) and IC₅₀ values of the complexes.

Complexes	Kb (M^−1)	IC50 (µM)
[Cu(SPF)(A^1)Cl].5H2O (1)	0.725 × 10^4	0.862
[Cu(SPF)(A^2)Cl].5H2O (2)	0.793 × 10^4	1.058
[Cu(SPF)(A^3)Cl].5H2O (3)	1.009 × 10^4	1.354
[Cu(PFL)(A^1)Cl].5H2O (4)	0.689 × 10^4	0.781
[Cu(PFL)(A^2)Cl].5H2O (5)	0.721 × 10^4	1.139
[Cu(PFL)(A^3)Cl].5H2O (6)	9.873 × 10^4	1.296

Figure 5.  Electronic absorption spectra of [Cu(SPF)(A¹)Cl].5H₂O in phosphate buffer (Na₂HPO₄/NaH₂PO₄, pH 7.2) in the absence and presence of increasing amount of DNA. The [Cu] complex = 10 μM; [DNA] = 0–150 μM. The incubation period is 30 min at 37°C. Inset: Plot of [DNA]/(ϵ_a– ϵ_f) vs. [DNA]. Arrow shows the absorbance change upon increasing DNA concentrations.

Figure 6.  Effect on relative viscosity of DNA under the influence of increasing amount of complexes at 27°C ± 0.1°C in phosphate buffer (Na₂HPO₄ /NaH₂PO₄, pH 7.2).

Table 6.  Complex mediated DNA cleavage data by gel electrophoresis.

Lane No	Compound	Form ISC	Form IIOC	Form IIILC
1	Control	83	17	-
2	CuCl2·2H2O	82	18	-
3	Sparfloxacin	58	42	-
4	Pefloxacin	54	46	-
5	[Cu(SPF)(A^1)Cl].5H2O (1)	11	75	14
6	[Cu(SPF)(A^2)Cl].5H2O (2)	14	65	21
7	[Cu(SPF)(A^3)Cl].5H2O (3)	24	63	13
8	[Cu(PFL)(A^1)Cl].5H2O (4)	19	69	12
9	[Cu(PFL)(A^2)Cl].5H2O (5)	12	77	11
10	[Cu(PFL)(A^3)Cl].5H2O (6)	11	71	18

Figure 7.  Photogenic view of cleavage of pUC19 DNA (300 µg/mL) with series of copper(II) complexes (200 µM) using 1% agarose gel containing 0.5 µg/mL ethidium bromide. All reactions were incubated in TE buffer (pH 8) in a final volume of 15 µL, for 24 h. at 37°C. Lane 1, DNA control; Lane 2, CuCl₂·2H₂O; Lane 3, Sparfloxacin; Lane 4, Pefloxacin; Lane 5, [Cu(SPF)(A¹)Cl].5H₂O; Lane 6, [Cu(SPF)(A²)Cl].5H₂O; Lane 7, [Cu(SPF)(A³)Cl].5H₂O; Lane 8, [Cu(PFL)(A¹)Cl].5H₂O; Lane 9, [Cu(PFL)(A²)Cl].5H₂O; Lane 10, [Cu(PFL)(A³)Cl].5H₂O.

Figure 8.  Absorbance values (Abs₅₆₀) as a function of time (t) plotted for varying concentration of complex 1 from 0.25 µM to 3 µM for produced a good straight line are observed.

Figure 9.  Plot of percentage of inhibiting NBT reduction with an increase in the concentration of complex 1.

de Souza MVN, de Almeida MV, da Silva AD, Couri MRC. Biological activity and synthetic methodologies for the preparation of fluoroquinolones, a class of potent antibacterial agents. Curr Med Chem 2003;10:21–39.

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