Insights into Structure and Biological Activity of Copper(II) and Zinc(II) Complexes with Triazolopyrimidine Ligands

Abstract

In an attempt to increase the biological activity of the 1,2,4-triazolo[1,5-a]pyrimidine scaffold through complexation with essential metal ions, the complexes trans-[Cu(mptp)₂Cl₂] (1), [Zn(mptp)Cl₂(DMSO)] (2) (mptp: 5-methyl-7-phenyl-1,2,4-triazolo[1,5-a]pyrimidine), [Cu₂(dmtp)₄Cl₄]·2H₂O (3) and [Zn(dmtp)₂Cl₂] (4) (dmtp: 5,7-dimethyl-1,2,4-triazolo[1,5-a]pyrimidine), were synthesized and characterized as new antiproliferative and antimicrobial species. Both complexes (1) and (2) crystallize in the P2₁/n monoclinic space group, with the tetrahedral surroundings generating a square-planar stereochemistry in the Cu(II) complex and a tetrahedral stereochemistry in the Zn(II) species. The mononuclear units are interconnected in a supramolecular network through π-π interactions between the pyrimidine moiety and the phenyl ring in (1) while supramolecular chains resulting from C-H∙∙∙π interactions were observed in (2). All complexes exhibit an antiproliferative effect against B16 tumor cells and improved antibacterial and antifungal activities compared to the free ligands. Complex (3) displays the best antimicrobial activity against all four tested strains, both in the planktonic and biofilm-embedded states, which can be correlated to its stronger DNA-binding and nuclease-activity traits.

Keywords: 1,2,4-triazolo[1,5-a]pyrimidine; biofilm; complex; cytotoxicity; metallonuclease activity.

Figure 1

The structure for 1,2,4-triazolo[1,5-a]pyrimidine derivatives described in the paper together with IUPAC ring-numbering system.

Scheme 1

The synthesis route of trans-[Cu(mptp)₂Cl₂] (1) and [Zn(mptp)Cl₂(DMSO)] (2).

Scheme 2

The synthesis route of [Cu₂(dmtp)₄Cl₄]·2H₂O (3) and [Zn(dmtp)₂Cl₂] (4).

Figure 2

The crystal structure for (1) with atoms labeled (a) and the supramolecular chains in compound (1) (b).

Figure 3

Crystal structure of (2): view of the asymmetric unit (a) and supramolecular chains resulting from C-H∙∙∙π interactions (b).

Figure 4

Powder EPR spectra of the complexes (1) and (3) at room temperature (a), EPR spectra of a single crystal of complex (1) with B₀ perpendicular to the ca (b), ab (c), and bc (d) planes, with rotation along the c, a, and b axes.

Figure 5

EPR spectra of complexes (1) and (3) in 10 mM DMSO solution: freshly prepared (a), after one week (b), and at temperatures ranging from 260 to 296 K for (1) (c) and (3) (d).

Figure 6

Cyclic voltammograms of copper complex (1) (a) and complex (3) (b), cyclic voltammograms of [Cu(DMSO)₆]Cl₂ (red line), ligand (blue line), complex ₍green line), 1 mM in DMSO, (0.1 M Bu₄NClO₄; scan rate: 0.050 V/s, working electrode, platinum disk, reference electrode, Ag/AgCl (0.1 M Bu₄NClO₄ in DMSO).

Figure 7

Cyclic voltammograms of zinc complex (2) (a) and complex (4) (b), cyclic voltammograms of [Zn(DMSO)₄]Cl₂ (red line), ligand (blue line), complex (green line), 1 mM in DMSO (scan rate: 0.100 V/s, working electrode: glassy-carbon disk, reference electrode: Ag/AgCl (0.1 M Bu₄NClO₄ in DMSO)).

Figure 8

Cyclic voltammograms of the electrolyte solution, 0.1 M Bu₄NClO₄ in DMSO without de-aeration with argon (red line), complex (2) (green line) and complex (4) (blue line), complexes’ concentration 1 mM in DMSO without de-aeration (scan rate: 0.100 V/s, working electrode: glassy-carbon disk: reference electrode, Ag/AgCl (0.1 M Bu₄NClO₄ in DMSO)).

Figure 9

The cytotoxic effect of the compounds (1)—(A), (2)—(B), (3)—(C) and (4)—(D) on B16 cells evaluated after 24 and 48 h (each value represents the mean ± SD).

Figure 10

Cell morphology of B16 cells: control cells (A), treated with 25 µM (B) and 75 µM (C) compound (1), treated with 25 µM (D) and 75 µM (E) compound (2), treated with 25 µM (F) and 75 µM (G) compound (3) and treated with 25 µM (H) and 75 µM (I) compound (4) for 24 h. Cell nuclei are stained with Hoechst 33342 (blue), and actin filaments are stained with Phalloidin-FITC (green). The scale bar is 20 μm and the same for all images.

Figure 10

Cell morphology of B16 cells: control cells (A), treated with 25 µM (B) and 75 µM (C) compound (1), treated with 25 µM (D) and 75 µM (E) compound (2), treated with 25 µM (F) and 75 µM (G) compound (3) and treated with 25 µM (H) and 75 µM (I) compound (4) for 24 h. Cell nuclei are stained with Hoechst 33342 (blue), and actin filaments are stained with Phalloidin-FITC (green). The scale bar is 20 μm and the same for all images.

Figure 11

EPR spectra of complex (1) (a) and (3) (b) interacting with KO₂ and H₂O₂ as O₂⁻ and OH⁻ radical sources, respectively.

Figure 12

Fluorescence of λ-DNA/EB in the absence (black curves, dotted line) or in the presence of increasing concentrations of mptp (a); complex (1) (b); complex (2) (c); dmtp (d); complex (3) (e); complex (4) (f). Arrows indicate the quenching of the fluorescence and the increase in compound concentrations. Concentrations used: λ-DNA, 3 μM; EB, 1 μM; mptp, dmtp, (1)–(4), 1–10 μM. Inset: I₀/I versus [quencher].

Figure 13

Gel electrophoresis image of pUC19 (100 ng/μL) after incubation for 1 h at 37 °C with the indicated compounds. (a) Lane 1: pUC19 alone; Lanes 2–7: pUC19 + 5 μM of indicated compounds. (b) Effect of (3) concentration on the relaxation of PUC19. SC, supercoiled; NC, nicked-circular.