Synthesis, characterization and DNA binding of magnesium–ciprofloxacin (cfH) complex [Mg(cf)₂] · 2.5H₂O

Abstract

Interactions of the tested systems (title compound [Mg(cf)₂] · 2.5H₂O (1), ciprofloxacin (cfH) and ciprofloxacin in the mixture with MgCl₂), with single and double stranded calf thymus DNA, poly[d(AT)] · poly[d(AT)] and poly[d(GC)] · poly[d(GC)] were studied by UV-spectrophotometric (melting curves) and fluorescence emission measurements. Pronounced quenching of ciprofloxacin’s fluorescence intensity has been observed for all the tested compounds after titration with various GC containing DNA molecules. It seems probable that quenching originates in the electron transfer from guanine to the photo-excited fluoroquinolone. The UV-spectrophotometric results obtained for 1 are substantially different from the other solutions and the biggest differences were observed for GC containing DNAs. Solution of 1 provokes a large thermal destabilization of poly[d(GC)] · poly[d(GC)]. This process is irreversible which suggests that the species present in solution of 1 alone inhibit re-annealing by associating irreversibly with the single strands. We have realized that aqueous solutions of 1 are colloidal and we propose that colloidal particles are involved in specific binding to GC containing sequences, most probably in the major groove of DNA.

Introduction

Quinolones are nowadays clinically the most successful synthetic antibacterial agents [1], [2], [3]. Fluoroquinolone member ciprofloxacin (cfH) (Scheme 1) is regularly found among the top 100 most frequently prescribed drugs in North America.

The emergence of drug-resistant bacteria is a growing problem also for quinolones, but maybe a new finding that blocking a protease called LexA could stop bacteria from evolving resistance to some antibiotics (including quinolones) could help to extend their use in the future [4].

Exact mechanism of quinolone action is not yet fully understood and recent review of the most important and sometimes contradictory results was given by Mitscher [1]. However, it is generally accepted that the quinolones target the bacterial enzyme gyrase–DNA complex which is responsible for the supercoiling of bacterial DNA [1].

It is known that nowadays several pharmaceutical companies have left the antibiotic discovery field and are much more interested in the more profitable areas of chronic diseases [5], [6]. Therefore it is crucial to understand the molecular mode of action of existing drugs which could help us to exploit them even more efficiently in the unpredictable and never ending battle between bacteria and mankind.

The interactions between metal ions and quinolones have been extensively studied for a long time and a review of metal complexes from this group was given [7]. Generally, quinolones coordinate to the metal as chelates through ring carbonyl and carboxylate oxygen atoms to form discrete molecules. In strongly acidic conditions quinolones are protonated and appear as cations in the metal complex compounds. It was also found that in the basic media the quinolones bearing a piperazinyl ring in position 7 (typical representatives ciprofloxacin (cfH), norfloxacin (nfH), pefloxacin), could also form complexes where terminal piperazinyl nitrogen is involved in the coordination to the metal [8], [9], [10], [11], [12], [13].

Up to now only few magnesium–quinolone crystal structures have appeared in the literature though it is well known that magnesium ions are important for the activity of these drugs [13], [14], [15]. The exact role of magnesium ions in the interaction between quinolone, DNA and DNA–gyrase is also not known yet and several models were proposed to explain the mechanism of action [16], [17], [18], [19].

In our previous work we have studied the interaction of cfH with calf thymus DNA, synthetic oligonucleotides and polynucleotides both in the presence and in the absence of metal ions (Cu²⁺, Mg²⁺) [20], [21], [22]. Our further aim was to prepare new magnesium–quinolone complex(es) and to study their interaction with DNA. Our efforts focused on the complexation in the vicinity of neutral pH that is comparable to the conditions found in the cells. At such pH a 2D molecular square-grid complex [Mg(cf)₂] · 2.5H₂O (1) was isolated by a hydrothermal reaction. Just before our paper was submitted we become aware of the work of Xiao et al. [13] who prepared and determined the crystal structures of 14 new metal–ciprofloxacin complexes. Among these also a magnesium complex [Mg(cf)₂] · 2.5H₂O was reported which is analogous to our title complex. In this complex two anionic molecules of ciprofloxacin are coordinated to magnesium ion through ring carbonyl and one of the carboxylate oxygens. The axial positions are occupied by two terminal nitrogen atoms of the piperazinyl residue resulting in the formation of a 2D metal-based molecular square grid. Water molecules are present in the channels that form in the structure. The main difference between crystallographic procedures employed by our group and that of Xiao et al. [13] was the temperature of the measurement. Due to the problem of the water disorder our crystal data were collected at low temperature (150 K), in contrast to Xiao et al. [13] who collected the data at 293 K. Unfortunately free mobility of water molecules in the channels prevents exact determination even at low temperature. Our low temperature data for compound [Mg(cf)₂] · 2.5H₂O have been deposited with the Cambridge Crystallographic Data Centre as supplementary publication no. CCDC 297483. Our results were partly reported before [23], [24] but there are still some important differences from synthetic and crystallographic view (see Section 3, Supplementary material) which we would like to stress in this paper. From the beginning our main goal was a biological relevance and not a physico-chemical characterization of the magnesium complex. This paper reports the comparison of the interactions between the tested systems (title compound 1, ciprofloxacin and ciprofloxacin in the mixture with MgCl₂), with single and double stranded calf thymus DNA, poly [d(AT)] · poly[d(AT)] and poly[d(GC)] · poly[d(GC)]. Understanding the interactions between studied compounds and DNA may help to elucidate the mechanism of action of this important class of antibacterial agents, and may ultimately lead to the design of better, more potent antibacterial agents.