Introduction

Metallopharmaceuticals playing a significant role in therapeutic and diagnostic medicine, the discovery and development of new metallodrugs remain an ever growing area of research in medicinal inorganic chemistry [1–3].

Imidazole and other azole ring systems are structural features of many drugs and natural products [4, 5]. Azoles are typically susceptible to N-alkylation, forming azolium cations [3]. Many of the latter undergo deprotonation of a ring C–H R to the nitrogen, generating an azolidene carbine [6]. In at least one case, that of thiamine (vitamin B1), such a carbene is thought to be involved in the biological function of the molecule [6–9]. In turn, the capacity of azolidene carbenes to function as ligands in transition-metal complexes is well-established [10, 11]. Consequently, biologically relevant molecules containing azoles are candidates as carbine sources for metal complexation. We consider the utilization of drugs or natural products in this role to have a potential and useful approach in the development of new compounds used in medicinal and biochemical applications. Further, since N-methylation is a common metabolic event [12–14]. Studies of such molecules may ultimately provide new insights into possible modes of metal-azole interactions in biological systems [15–17].

Metallic silver, silver salts, and silver complexes have been used in a variety of applications like water purification, wound management, eye-drops, anti-infective coatings in medical devices and in burn treatment because they have potent antimicrobial properties but with low human toxicity [18–24]. Also they represent novel group of antifungal agents with potential applications for the control of fungal infections. Treatment of fungal cells with the Cu (II) and Ag (I) complexes resulted in a reduced amount of ergosterol in the cell membrane and subsequent increase in its permeability [25–27].

However, in this study metal complexes of Mn(II), Cr(III), Fe(III), Co(II), Ni(II), Cu(II), and Zn (II) transition metals with MCNZ drug molecule were prepared.

This new complexes are linked a drug-based ligand to a metal via a metal-nitrogen bond, and they complement a small family of antifungal drug-metal conjugates possessing metal-nitrogen linkages [28].

The solid complexes are characterized using different physico-chemical methods like elemental analyses (C, H, N, Cl, and metal content), IR, magnetic moment and reflectance spectra, and thermal analyses (TGA and DTA). Biological activities of the complexes are studied. The structure of the drug is given in Fig. 1.

Fig. 1
figure 1

Structure of MCNZ drug

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Experimental section

Materials

All chemicals used were of the analytical reagent grade (AR), and of highest purity available. They included miconazole (Union P.Goup); CuCl2·2H2O (Prolabo); CoCl2·6H2O and NiCl2·6H2O (BDH); ZnCl2·2H2O (Ubichem), CrCl3·6H2O (Sigma); MnCl2·2H2O, and FeCl3·6H2O (Prolabo). Organic solvent used was absolute ethyl alcohol. De-ionized water collected from all glass equipments was usually used in all preparations.

Instruments

Elemental microanalyses of the separated solid chelates for C, H, N, and Cl were performed at the Microanalytical Center, Cairo University. Infrared spectra were recorded on a Perkin-Elmer FT–IR type 1,650 spectrophotometer in wavenumber region 4000–400 cm−1. The spectra were recorded as KBr pellets. The mass spectrum of the drug under investigation was recorded by EI technique at 70 eV using MS-5988 GS–MS Hewlett-Packard instrument at the Microanalytical Center, Cairo University. The thermal analyses (TG, DTG, and DTA) analyses were carried out in dynamic nitrogen atmosphere (20 mL min−1) with a heating rate of 10 °C min−1 using Shimadzu TG-60H and DTA-60H thermal analyzers. The solid reflectance spectra were measured on a Shimadzu 3101pc spectrophotometer. The molar magnetic susceptibility was measured on powdered samples using the Faraday method. The diamagnetic corrections were made by Pascal’s constant and Hg [Co (SCN)4] was used as a calibrant.

Synthesis of metal complexes

Hot ethanolic solution (60 °C) of the appropriate metal chloride (1 mmol) in an ethanol–water mixture (1:1, 25 mL) was added to 25 mL of MCNZ (0.5 g, 2 mmol) hot ethanolic solution (60 °C). The resulting mixture was stirred under reflux for 1 h whereupon the complexes precipitated. They were collected by filtration, purified by washing several times with a 1:1 ethanol: water mixture and petroleum ether. The analytical data for C, H, N, and Cl were repeated twice.

Biological activity

The ligand and its corresponding metal complexes were screened in vitro for their antimicrobial activities against Bacterial strains (Pseudomnas aeruginosa, Staphylococcus aureous, Bacillus subtilis, Escherichia coli) and fungi strains (Aspergillus fumigates, Penicillium italicum, Synephalastrum racemosum, and Candida albicans) using agar well diffusion method. The samples were dissolved in DMSO to make a concentration of 100 μg/mL ligand and complexes and 2 mg/mL for standards. The inoculum (1 × 108 Cfu/mL) was added to molten agar and the media were shaken to disperse the microorganisms. Four millimeters diameter wells were punched in the agar with a sterile cork borer. 10 μl of the sample was introduced in the well. Antimicrobial activity was evaluated by measuring the diameter of inhibition zone in mm. The experiments were conducted in triplicate [29–31].

Results and discussion

The formation of metal complexes with organic compounds has long been recognized. However, the binary complexes of the cited drug with metal ions have not been studied yet, although they may be an area of interest. This is because they may affect the bioavailability of these drugs as certain metal ions were present in relatively appreciable concentration in biological fluids.

The different pathways of the possible fragments of MCNZ with its respective relative intensities (RI) are given in Scheme 1. Fragment at m/z = 414 (RI = 100%) represents the base peak of MCNZ. The order molecular ion peaks appeared in the mass spectrum is attributed to the fragmentation obtained from the rupture of different bonds inside the drug molecule.

Scheme 1
scheme 1

Mass fragmentation pattern of MCNZ drug

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Composition and structures of metal complexes

The metal ions and MCNZ ligand was condensed in the ratio of 1:2 for the preparation of mononuclear complexes. The isolated solid complexes of Cr(III), Mn(II), Fe(III), Co (II), Ni (II), Cu (II), and Zn (II) were investigated by elemental analyses (C, H, N, Cl, and metal content), IR, magnetic moment studies, molar conductance, and thermal analyses (TG and DTA). The results of elemental analyses listed in Table 1 suggest that they have the composition [MCl2 (H2O)2(L)2]·yH2O (where y = 1 for M = Mn(II) or y = 2 for M = Cu(II), Co(II)), and y = 3 for M = Ni(II), Zn(II), and [MCl2(H2O)2(L)2]Cl·3H2O (where M = Cr(III) and Fe(III)). The results obtained are agreed well with the proposed formulae.

Table 1 Analytical and physical data of MCNZ metal complexes
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Molar conductivity measurements

The conductance measures the ability of an aqueous solution to carry an electric current. It is concluded from the results that the complexes of the bivalent metal ions were found to have molar conductance values of 13–23 Ω−1 mol−1 cm2, which indicates the non-ionic nature of these complexes and they are considered as non-electrolytes [32]. While for Cr(III) and Fe(III) complexes, the molar conductance values indicate the ionic nature of these complexes and they are of the type 1:1 electrolytes.

IR spectral studies and mode of bonding

The IR spectral data of the MCNZ drug and its metal complexes are listed in Table 2. The position and/or the intensities of these bands are expected to be changed upon chelation. These characteristic bands are listed in Table 2. Upon comparison it is found that the band located at 1631 cm−1 is assigned to the ν(C=N) stretching vibration of imidazole nitrogen for MCNZ drug [32]. Coordination of the imidazole nitrogen to the metal ion is indicated by a 3–9 and 4–23 cm−1 shift to lower and higher wave numbers, respectively. The band located at 1088 cm−1 is assigned to the ν(C–O–C) in the free drug. This band remains intact in solid complexes, indicating its non-involvement in coordination to the metal ions. The bands located at 3376–3550 and 907–964 cm−1 are assigned to the stretching vibrations of the hydrated and coordinated water molecules, respectively. The structural formula of the drug under investigation shows that it is promising in forming metal chelates through the N of the imidazole ring. According to IR data, it is concluded that MCNZ drug behaves as a neutral unidentate ligand coordinated to the metal ions via the imidazole N.

Table 2 IR spectral data (4000–400 cm−1) of the MCNZ drug and its metal complexes
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Magnetic susceptibility and electronic spectra measurements

From the diffused reflectance spectrum, it is observed that, the Fe(III) complex exhibits band at 20820 cm−1, which may be assigned to the 6A1g → T2g(G) transition in octahedral geometry of the complexes. The 6A1g → 5T1g transition appears to be split into two bands at 16950 and 14650 cm−1. The observed magnetic moment of the Fe(III) complex is 5.82 BM [33]. Thus, the complex formed has the octahedral geometry. The spectrum shows also band at 27690 cm−1 which may attributed to ligand to metal charge transfer.

The diffused reflectance spectrum of the Mn(II) complex displays four medium intensity bands at 15562, 18530, 21739, and 25340 cm−1 assignable to 6A1g → 4T1g(4G), 6A1g → 4Eg, 6A1g → 4Eg(4D), and 6A1g → 4T1g(4P) transitions [33, 34]. The magnetic moment recorded at room temperature is found to be 5.61 BM corresponding to five unpaired electrons in octahedral geometry [35]. The diffused reflectance of the Co(II) complex with the general formula [CoCl2(L)2(H2O)2]·2H2O, gives three bands at 15550, 17820, and 19683 cm−1, which are assigned to the transitions 4T1g (F) → 4T2g (F) (ν1), 4T1g (F) → 4A2g (F) (ν2) and 4T1g (F) → 4T2g (P) (ν3), respectively, suggesting that the Co(II) complex has octahedral geometry [36, 37].

The diffused reflectance spectra of the Cr(III) chelate exhibit bands at 21341–23730 which may be assigned to the 6A1g → T2g(G) transition in octahedral geometry of the complex. The observed magnetic moments of Cr(III) complex are 5.16, 5.47 BM The spectra show also bands at 25281–27645 cm−1 which may be attributed to ligand to metal charge transfer [37].

The diffused reflectance of the Ni (II) complex (beside the π–π* and n–π* bands of free drug) display three bands at ν1: 15770 cm−1: 3A2g → 3T2g; ν2: 19250 cm−1: 3A2g → 3T1g (F), and ν3: 22350 cm−1: 3A2g → 3T1g(P). The spectrum shows also a band at 25390 cm−1 which may attributed to ligand to metal charge transfer.

The magnetic moment values for Co (II) and Ni (II) complexes indicate the high-spin octahedral nature of the complexes [34–37].

The reflectance spectrum of Cu (II) complex exhibits a band at 14777 cm−1 which is assigned to the transition 2Eg/2T2g in an octahedral geometry [34–37]. The magnetic moment of 2.05 BM falls within the range normally observed for octahedral Cu (II) complex. A moderately intense peak observed in the range 21555 cm−1 is due to ligand to metal charge transfer transition.

The Zn (II) complex is diamagnetic and an octahedral geometry was proposed based on its empirical formula.

Thermal analyses (TG, DTG, and DTA)

The TG curve of MCNZ (Fig. 2a) exhibits an estimated mass loss of 24.82% (calcd. 24.85%) within the temperature range 50–250 °C, which may be attributed to the liberation of C3H3N2 and HCl molecules as gases. In the second and third stages within the temperature range 250–700 °C, the remaining part; C15H10Cl3O molecule is lost with an estimated mass loss of 75.18% (calcd. 75.12%) with a complete decomposition as CO, CO2, NO, NH3, etc., gases.

Fig. 2
figure 2

Thermal analyses of (a) MCNZ drug (b) Fe(III) (c) Cu(II) (d) Co(II) (e) Mn(II) and (f) Ni(II) complexes

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The thermal curve of the [FeCl2 (L)2(H2O)2]Cl·3H2O complex (Fig. 2b) shows four decomposition steps within the temperature range 30–700 °C. The first step of decomposition is found to be within the temperature range 30–100 °C which corresponds to the loss of water molecules of hydration with a mass loss of 4.39% (calcd. 4.98%). The energy of activation is found to be 31.02 kJ mol−1. In the second stage within the temperature range 100–230 °C, the weight loss found may the decomposition of 3HCl molecules with a mass loss of 10.61% (calcd. 10.10%). The subsequent steps (230–700 °C) correspond to the removal of the two ligand molecules and ½H2 leaving metal oxide as a residue. The overall weight loss amounts to 91.39% (calcd. 91.09%).

The TG curves of the Cu(II) (Fig. 2c) and Co(II) (Fig. 2d) complexes show from three to four decomposition steps within the temperature range of 30–800 °C. The stages at 30–400 °C of the Cu(II) complex or 30–450 °C for Co(II) complex correspond to the loss of 3H2O, 2HCl, and C14H8Cl4O molecules. The energy of activation for these steps is 54.80 and 86.03 and 12.85 kJ mol−1 for Cu(II) and Co(II) complexes, respectively. While the subsequent stages involve the loss of C22H20Cl4N4O molecule (as shown in Table 3), with a mass loss of 48.18% (calcd. 47.95%) and 48.03% (calcd. 48.16%) for Cu(II) and Co(II) complexes, respectively. The overall weight loss amounts to 93.04% (calcd. 92.34%) and 92.69% (calcd. 92.72%) for Cu(II) and Co(II) complexes, respectively.

Table 3 Thermal analyses (TG, DTG, and DTA) data of MCNZ drug and its metal complexes
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Mn(II) complex is thermally decomposed in four decomposition steps, Fig. 2e, within the temperature range 30–700 °C. The first decomposition step with an estimated mass loss of 22.58% (calcd. 22.75%) within the temperature range 30–220 °C may be attributed to the decomposition of 2HCl, 2H2O, and C4H2Cl2 molecules. The activation energy is 45.60 kJ mol−1. The remaining decomposition steps found within the temperature range 220–700 °C with an estimated mass loss of 41.54 (calcd. 41.34) and 29.19% (calcd. = 28.79%) are reasonably accounted for the removal of C20H19Cl2 N4O2 and C12H7Cl4 molecules as gases, respectively.

The TG curve of the Ni(II) (Fig. 2f) complex represents four decomposition steps as shown in Table 3. The first two step of decomposition within the temperature range 25–250 °C correspond to the loss of 4H2O, 2HCl, and C6H3Cl2 molecules with a mass loss of 27.48% (calcd. 27.66%). The energy of activation for this dehydration steps are 33.66 and 49.901 kJ mol−1. The third stage within the temperature range 250–400 °C which may be attributed to the loss of C15H14Cl2N4O molecule with a mass loss of 32.04% (calcd. 32.40%). The subsequent step (400–700 °C) corresponds to the removal of C15H11Cl4O molecule leaving metal oxide as a residue. The overall weight loss amounts to 93.12% (calcd. 93.23%).

The DTA results are listed in Table 3 and represented graphically in Fig. 2. The appearance of many exothermic and endothermic peaks in the DTA (Table 3) can be attributed to the gaseous decomposition of the water, anions, and ligand molecules during decomposition process of the complexes [38].

Biological activity

Since the use of MCNZ drug as effective antifungal drug, the interest toward transition metal complexes containing N-donor ligands has increased in order to obtain metal-based drugs exhibiting a high biological activity together with a reduced toxicity. In this respect, in vitro biological activity of MCNZ drug with its transition metal complexes have been investigated. The data obtained shows that MCNZ drug under investigation and its metal complexes have the capacity of inhibiting the metabolic growth of the investigated bacteria and fungi to different extent. The size of the inhibition zone depends upon the culture medium, incubation conditions, rate of diffusion and the concentration of the antibacterial agent. The activities of all the tested complexes may be explained on the basis of chelation theory; chelation reduces the polarity of the metal atom mainly because of partial sharing of its positive charge with the donor groups and possible π electron delocalization within the whole chelate ring. Also, chelation increases the lipophilic nature of the central atom which subsequently favors its permeation through the lipid layer of the cell membrane.

In testing the antibacterial and antifungal activity of MCNZ drug and its metal complexes, more than one test organism is used to increase the chance of detecting antibiotic principles in tested materials. The sensitivity of a microorganism to antibiotics and other antimicrobial agents is determined by the assay plates which incubated at 28 °C for 2 days for yeasts and at 37 °C for 1 day for bacteria. All of the tested compounds show a remarkable biological activity against different types of fungi (Aspergillus fumigatus, Penicillium italicum, Syncephalastrum racemosum, and Candida albicans), Gram-positive bacteria (Staphylococcus aureus, Pseudomonas aeruginosa), and Gram-negative bacteria (Bacillus subtilis, Escherichia coli). The data are listed in Table 4. It is clear from the obtained results that:

Table 4 The antibacterial and antifungal activity of MCNZ drug and its metal complexes
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  1. (a)

    Using A. fumigatus and P. italicum fungi: The data listed in Table 4 shows that Cr(III) and Fe(III) complexes have the higher biological activity than that of the free drug while the Cu(II), Co(II), Mn(II), Ni(II), and Zn(II) drug complexes show no biological activity.

  2. (b)

    Using S. racemosum fungus: It is clear from Table 4 that MCNZ and its metal complexes have no antifungal activity.

  3. (c)

    Using Candida albican fungus: MCNZ and its metal complexes show moderate effect on this kind of fungus, The inhibition values are ranged from 0.1 to 1.0 mm beyond control. The data listed in Table 4 illustrate that the antifungal activity is found to follow the order: Mn(II) = Ni(II) > MCNZ > Co(II) = Cu(II) > Cr(III), while Zn(II) and Fe(III) complexes are found to have no antifungal activity.

  4. (d)

    Using S. aureus (G ± bacteria): It is obvious from the data listed Table 4 that MCNZ and its metal complexes have the same antibacterial activity.

  5. (e)

    Using P. aeruginosa bacteria: The results also show that metal complexes of MCNZ drugs have no activity, but Cu(II), Co(II), Ni(II), Zn(II), and Fe(III) complexes of MCNZ have bacterial activity at higher concentrations (5 mg L−1) only.

  6. (f)

    Using B. subtilis bacteria: MCNZ and its metal complexes are found to have antibacterial activity against B. subtillis bacteria (an example of G− negative bacteria), and the activity is found to have the order: Fe(III) > Ni(II) > MCNZ = Cu(II) = Co(II) Mn(II) = Zn(II) = Cr(III).

  7. (g)

    Using E. coli bacteria

It is clear that Zn(II) complex of MCNZ drug only show antibacterial activity against E. Coli (Table 4).

Structural interpretation

The structures of the complexes of MCNZ drug with Cr(III), Mn(II), Fe(III), Co(II), Ni(II), Cu(II), and Zn(II) ions are confirmed by the elemental analyses, IR, molar conductance, magnetic, solid reflectance, mass, and thermal analyses (TG and DTA) data. Therefore, from the IR spectra, it is concluded that MCNZ drug behaves as a neutral undentate ligand coordinated to the metal ions via the imidazole-N. The molar conductance data, it is found that the bivalent complexes are non electrolytes while trivalent complexes are 1:1 electrolytes. On the basis of the above observations and from the magnetic and solid reflectance measurements, octahedral geometrical structure is suggested for the investigated complexes. As a general conclusion, the investigated MCNZ drug complexes structurs can be given as shown in Fig. 3.

Fig. 3
figure 3

Structures of metal complexes

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