Properties of Binuclear Rhodium(II) Complexes and Their Antibacterial Activity

Binuclear rhodium(II) complexes [Rh2Cl2(μ-OOCR)2(N-N)2], [Rh2(μ-OOCR)2(N-N)2(H2O)2](RCOO)2 and [Rh2Cl2(μ-OOCCH3)(terpy)2](H3O)Cl2.9H2O (R = H, Me, Bun, ph, PhCHOH; N-N = 2,2′-bipyridine (bpy), 1,10-phenanthroline (phen), 2,9-dimethyl-1,10-phenanthroline (dmp) and 6,7-dimethyl-2,3- di(2-pyridyl)quinoxaline (dmpq); terpy 2,2′:6′,2′′-terpyridine) have been synthesized and their structure and properties have been studied by electronic, IR and 1H NMR spectroscopy. Antibacterial activity of these complexes against Staphylococcus aureus and Escherichia coli has been investigated. The most active antibacterial agents against S. aureus were [Rh2(OOCPh)2(phen)2(H2O)2]2+, [Rh2(OOCPh)2(dmpq)2(H2O)2]2+, [Rh2(OOCBu)2(phen)2(H2O)2]2+ and [Rh2-(OOCBu)2(bpy)2(H2O)2]2+ which were considerably more active than the appropriate nitrogen ligands. The complexes show rather low activity against E. coli.

Recently it has been found [18] that [Rh2(OOCR)2(N-N)2 0)2 complexes show cytostatic activity for human oral carcinoma KB cell-line in vitro. Some the determined biological activities were in close relation with findings obtained on plant cell cultures [19], when synchronously cultivated green algae were exposed to Rh(ll) complexes. New metal compounds possessing antimicrobial activity are recently intensively investigated because of the increasing resistance of microbes against many antibiotics. It has been found that some platinum complexes possessing antitumor activity show also antibacterial properties. A further well defined series of metal complexes with antibacterial activity is that of rhodium(Ill) coordination compounds of formula trans-[RhX2(py)4]Y [20][21][22]. These complexes are more active against Gram-positive microorganisms than Gram-negative ones. However, the complexes [RhCI2(bpy)2] / are not active antibacterial agents. There is considerable interest in interactions of tetra-#-carboxylato-dirhodium(ll) complexes with nucleic acid bases because they function as antitumor agents against many types of tumors by inhibiting DNA or protein synthesis. The examination of the biological activity of these compounds in such living systems (i.e. tumor bearing animals) is time consuming and expensive and should be used at the last step of examination. The exposure of mammalian cells in cultures or microorganisms cultivated synchronously, or an examination of the antibacterial activity of metal compounds [19,23] give also interesting information about the general biological activity of the tested compounds. In this paper, we report on synthesis and properties of dimeric rhodium(ll) complexes with carboxylato and heterocyclic nitrogen ligands and their antibacterial activity. 1,10-Phenanthroline (phen), 2,9-dimethyl-l,10-phenanthroline (dmp), 2,2'-bipyridine (bpy), 2,2':6',2"-terpyridine (terpy) and 6,7-dimethyl-2,3-bis(2-pyridyl)quinoxaline (dmpq) were obtained from Aldrich and used without further purification. [Rh,,2(OOCCHOHPh)(.1269(dmP)2(H20)2](OOCCHOHPh)2(1).
A solution of [Rh2(OOCH-OHPh)4(H20)2 g, 0.15 mmol) and dmp (0.0625 g, 0.3 mmol) in 6 cm 3 of ethanol was heated at reflux with stirring for 2 h. The initial green colour of the solution was soon replaced by brown-red and then by deep green-blue. The deep green-blue rhodium (I) complexes are formed because rhodium(ll) dimers are reduced by ethanol. However, rhodium(I) compounds are very readily oxidized to red-brown dimeric rhodium(ll) complexes under an atmosphere of air.
They are stable under air atmosphere both in solid state and in solutions, readily soluble in water, methanol and ethanol and slightly soluble in higher alcohols, insoluble in diethyl ether and nonpolar solvents. Primary and secondary alcohols at elevated temperatures reduce these complexes giving rhodium(I) complexes, that are very readily reoxidized to rhodium(ll) compounds by means of air.
All complexes have been characterized by a combination of elemental analysis and IR, UV-VIS and H NMR spectroscopy. The data are collected in Tables 1-3.
The physicochemical measurements indicate that structure of the cationic complexes containing two bridging carboxylato ligands is analogous to that of Rh2CI2(RCOO)2(N-N)2 [2,4]      nm. This indicate that adenine is coordinating to rhodium via nitrogen atom. However, in the case of adenosine blue shift is much lower (ca. 2-5 nm) suggesting that this ligand is coordinating through oxygen atom or undergoes stacking owing to the r interaction between the purine moiety and a heteroaromatic amine bound with rhodium atom. The stacking of nucleobases and metal complexes is well known [29][30][31]. The IR spectra of complexes with phenanthrolines showed that the carboxylates are symmetrically bonded to the both rhodium atoms because differences between vas(coo) and vs(coo) are small ( Table 2).
The phenanthroline, 2,2'-bipyridine and 2,2':6',2"-terpyridine ligands in atoms. Ths s. proven by the presence only 4 signals of hydrogen atoms of heter,o, cyclic nitr,,ogen ligands in the 1H NMR spectra of phenanthroline complexes and signals of H5 + H5, H6 + H6 and multiplet for the other protons of 2,2':6',2"terpyridine ligand in complex 9 ( Table 3). The H NMR spectrum of terpy complex is consistent with the X-ray structure of this compound [26]. However, the H NMR spectra of complexes 1-4, and 13 consist of multiplets due to non-equivalent protons of the nitrogen ligands. Thus, the nitrogen ligands are coordinated asymmetrically in complexes containing chiral mandelato bridging groups and in acetato complex with dmpq. An unambiguous assignment of the spectra was possible only for the compounds 2 and 3. In the case of complexes 1,4 and 13 most of the signals were overlapped and therefore they could not be unambiguously assigned.   nc noncoordinating ion, c coordinating ligand, qnt quintet, sxt sextet The complexes 1-13 and the ligands (phen, drop, bpy, terpy and dmpq) were tested in vitro for antibacterial activity against Gram-positive Staphylococcus aureus 209 P (Oxford) and Gramnegative Escherichia coil ROW strains ( Table 4). The highest activity against Gram-positive bacteria axhibited complexes 2-6 and 10-12. Compounds 1 and 7 are much less active against S. aureus and all complexes show rather low activity against E. coil The complexes 2, 4, ,5 and 10-12 are considerably more active than the appropriate nitrogen ligands. However, the activity of complex 1 is lower than that of drop. The activity of 3 is almost the same as that for non-coordinated phen and lower than activity of 2. The complex 3 in the solid state contains chloro ligands coordinated along the Rh Rh axis, however in solutions in water and in alcohols the prevailing form is [Rh2CI{PhCH(OH)COO}2(phen)(H20)] + [2]. This indicate that complexes containing aqua molecules coordinated along Rh -Rh axis are more active than those with at least one chloro ligand substituted for HO. This is also confirmed by low activity of complex 9 with two chloro ligands bound along the Rh Rh axis [26]. In order to get more information about the nature of the inhibitory activity of the investigated compounds, we have tested the survival of S. aureus in YP medium containing different concentration of complexes 2, 4, ,5 and 6. Viable counts were determined after 4 and 24 h and the results are presented in Fig. 2 It is known that the bactericidal activity of some antibiotics and chemoterapeutic agents depend on the size of thepopulation of treated microorganisms. Therefore, we have determined the survival of S. aureus cultures over the population range 10 to 106 cells/cm 3 in the presence of ,5. The results presented in Table 5 indicate that the bactericidal activity of this compound depends on inoculum size. The bacteria are killed at the lowest concentration of ,5 (6.10 7 M) when the inoculum size is equal to or less than 10 ' cells/cm3, however, for the populations equal to or greater than 105 cells/cm3, the complex 5 is effective at the concentration 1.25.10 -6 M.  *determined on nutrient agar plates supplemented with tested rhodium complexes not tested Some of the antibacterial agents, e.g. penicillin, are active only against growing cells. Thefore, it was interesting to find out whether the investigated compounds are active only against growing or also resting cells. The dependence of the survival of S. aureus treated with compound ,5 on its concentration and time is presented in Table 6. It is evident that the complex ,5 shows a killing effect on cells unable to grow. Incubation in saline even without the rhodium complexes leads to a decrease in viable count of S. aureus but in the presence of the compound 5 the diminution of the percent of colony forming units is much more drastic and depends on concentration. Analogous investigations were also carried out for complex 4.  The results given in Table 7 indicate that compound 4 is also bactericide. This compound shows bactericidal activity against nongrowing cells suspended in saline ( Table 8). The activity of the compound 5 was also tested against some of clinical S. aureus strains chosen at random. From among of 19 strains only 1 was resistant to the complex ,5 in YP medium.