Synthesis of New Zirconium Magnetic Nanocomposite as a Bioactive Agent and Green Catalyst in the Four-Component Synthesis of a Novel Multi-Ring Compound Containing Pyrazole Derivatives

New nanocomposites containing zirconium were synthesized using microwave irradiation. Their structure was confirmed by vibrating sample magnetometer (VSM) curves, X-ray diffraction (XRD) patterns, scanning electron microscope (SEM) and transmission electron microscopy (TEM) images, Fourier transform infrared spectroscopy (FT-IR), and Brunauer–Emmett–Teller (BET) N2 adsorption/desorption isotherms. After the structure confirmation of the zirconium magnetic nanocomposite, the catalytic properties in the synthesis of pyrazole derivatives were investigated. Next, the biological activities of the zirconium magnetic nanocomposite, such as the antibacterial and antifungal activities, were investigated. The research results showed that the zirconium magnetic nanocomposite has high catalytic properties and can be used as a magnetic nanocatalyst for synthesizing heterocyclic compounds such as pyrazole derivatives in addition to having high biological properties. The unique properties of the nanoparticles can be attributed to their synthesis method and microwave radiation.


Introduction
Cyclic organic compounds withwith at least one heteroatom, such as nitrogen, sulfur, and phosphorus, are called heterocycles. Heterocycles have many biological properties. There are heterocyclic compounds in the structures of many drugs. So far, biological properties such as antibacterial, antifungal, anticancer, and antioxidant properties of heterocyclic compounds containing nitrogen, sulfur, and phosphorus have been reported [1][2][3][4][5][6][7]. One of criticalheterocyclic compounds' critical applications is their use as ligands in complexes. The use of heterocycles as ligands makes the final product retain the biological properties of the heterocycle and the metal, thus having high biological properties. There have been reports of using heterocycles as ligands and synthesizing novel complexes containing various metals such as Cr, Mo, W, gold, and silver. These have unique properties, including anticancer properties [8][9][10][11][12].

Solvents and Raw Materials
The high-purity solvents and raw materials used in this study were purchased from Merck and Sigma-Aldrich. The Fe 3 O 4 nanostructures were prepared from Sigma-Aldrich. No purification of the raw materials was carried out (Merck KGaA, St. Louis, MO, USA).

Zirconium Magnetic Nanocomposite Synthesis
For the zirconium magnetic nanocomposite synthesis, Fe 3 O 4 nanoparticles (2 mmol), dipicolinic acid (4 mmol), and ZrCl 4 (2 mmol) were added to 30 mL double-distilled water and stirred at 80 • C. After 10 min, the solution was put into a microwave and irradiated at a microwave power of 450 W at room temperature. After 10 min, the mixture cooled (room temperature), and the desired product was isolated using an external magnet. The synthesized zirconium magnetic nanocomposite was washed several times with a mix of double-distilled water and ethanol and dried at an ambient temperature.

Zirconium Magnetic Nanocomposite Antimicrobial Activity
To measure the MIC, MBC, and MFC, a concentration of 1-2048 mg/mL of zirconium magnetic nanocomposite and the drug were prepared. The Clinical and Laboratory Standards Institute (CLSI) guidelines (M07-A9, M26-A, M27-A2) were used for the zirconium magnetic nanocomposite antimicrobial activity. Based on the reported methods, relevant tests on the desired Gram-positive, Gram-negative species, and desired fungal species were performed [34][35][36].
The magnetic saturation of the zirconium magnetic nanocomposite, as shown in the VSM curve in Figure 1, was 0.014 emu/g.
The magnetic property of the zirconium magnetic nanocomposite was compared with the magnetic property of the Fe 3 O 4 nanoparticles. According to previous reports, the saturation value of the Fe 3 O 4 nanoparticles was 0.055 emu/g [2]. The decrease in the magnetic saturation of the zirconium magnetic nanocomposite shows that the Fe 3 O 4 nanoparticles were covered in groups. The magnetic property of the zirconium magnetic nanocomposite was compared with the magnetic property of the Fe3O4 nanoparticles. According to previous reports, the saturation value of the Fe3O4 nanoparticles was 0.055 emu/g [2]. The decrease in the magnetic saturation of the zirconium magnetic nanocomposite shows that the Fe3O4 nanoparticles were covered in groups.
The XRD pattern of the zirconium magnetic nanocomposite ( Figure 2) confirmed the crystalline structure and the presence of Fe3O4 nanoparticles in the final product's structure [37]. The SEM and TEM images of the zirconium magnetic nanocomposite ( Figure 3) confirmed the uniformity of the structure and the morphology of the final product. In addition, the SEM and TEM images proved that the structure of the compound was in the nano-sized range.  The magnetic property of the zirconium magnetic nanocomposite was compared with the magnetic property of the Fe3O4 nanoparticles. According to previous reports, the saturation value of the Fe3O4 nanoparticles was 0.055 emu/g [2]. The decrease in the magnetic saturation of the zirconium magnetic nanocomposite shows that the Fe3O4 nanoparticles were covered in groups.
The XRD pattern of the zirconium magnetic nanocomposite ( Figure 2) confirmed the crystalline structure and the presence of Fe3O4 nanoparticles in the final product's structure [37]. The SEM and TEM images of the zirconium magnetic nanocomposite ( Figure 3) confirmed the uniformity of the structure and the morphology of the final product. In addition, the SEM and TEM images proved that the structure of the compound was in the nano-sized range. The SEM and TEM images of the zirconium magnetic nanocomposite ( Figure 3) confirmed the uniformity of the structure and the morphology of the final product. In addition, the SEM and TEM images proved that the structure of the compound was in the nano-sized range.  The FT-IR spectrum of the zirconium magnetic nanocomposite, as shown in Figure  4, proved the desired absorptions of the final product's structure. The FT-IR spectrum of the zirconium magnetic nanocomposite, as shown in Figure 4, proved the desired absorptions of the final product's structure.  The N2 adsorption/desorption isotherms of the zirconium magnetic nanocomposite were the fourth type of the classical isotherm series [39]. The specific surface area of the zirconium magnetic nanocomposite was about 1850 m 2 /g ( Figure 5). The N 2 adsorption/desorption isotherms of the zirconium magnetic nanocomposite were the fourth type of the classical isotherm series [39]. The specific surface area of the zirconium magnetic nanocomposite was about 1850 m 2 /g ( Figure 5). observed in the 3400 cm −1 .
The N2 adsorption/desorption isotherms of the zirconium magnetic nanocom were the fourth type of the classical isotherm series [39]. The specific surface area zirconium magnetic nanocomposite was about 1850 m 2 /g ( Figure 5). As an overall finding, it can be stated that the synthetic compound had ma properties and could be easily separated in catalytic reactions. The desired elemen functional groups were observed in the structure of the synthesized compound. Th thesis method and microwave radiation caused uniform morphology and nano-siz ticles. In addition, the synthesis method increased the specific surface area, which its use as an efficient catalyst and bioactive agent.
Based on the observations and spectral analysis, the following structure wa gested for the zirconium magnetic nanocomposite ( Figure 6). As an overall finding, it can be stated that the synthetic compound had magnetic properties and could be easily separated in catalytic reactions. The desired elements and functional groups were observed in the structure of the synthesized compound. The synthesis method and microwave radiation caused uniform morphology and nano-sized particles. In addition, the synthesis method increased the specific surface area, which led to its use as an efficient catalyst and bioactive agent.
Based on the observations and spectral analysis, the following structure was suggested for the zirconium magnetic nanocomposite ( Figure 6).

Results of four-Component Synthesis of Multi-Ring Compound Containing Pyrazole Using Zirconium Magnetic Nanocomposite
From the four-component reaction of the aromatic aldehyde derivatives, malononitrile, phenylhydrazine, and ethyl acetoacetate in the presence of the zirconium magnetic nanocomposite as a catalyst, 1,4-dihydropyrano[2,3-c]pyrazole-5-carbonitrile derivatives were synthesized (Scheme 1).
To synthesize the derivatives, we first optimized the reaction conditions, such as the solvent, amount of catalyst, and temperature. Different solvents, such as EtOH, EtOH H2O (1:1), MeOH, and CH3CN, were tested during optimization. Based on the obtained results, the highest efficiency was obtained using EtOH: H2O.
To optimize the catalyst amount, the reactions in amounts of 1-5 mg were tested. Based on the obtained results, high efficiency was observed in using 4 mg as the catalyst Finally, temperature optimization was performed, and the reaction at 50 °C had the highest yield. The optimization results are given in Table 1. To synthesize the derivatives, we first optimized the reaction conditions, such as the solvent, amount of catalyst, and temperature. Different solvents, such as EtOH, EtOH: H2O (1:1), MeOH, and CH3CN, were tested during optimization. Based on the obtained results, the highest efficiency was obtained using EtOH: H 2 O.
To optimize the catalyst amount, the reactions in amounts of 1-5 mg were tested. Based on the obtained results, high efficiency was observed in using 4 mg as the catalyst. Finally, temperature optimization was performed, and the reaction at 50 • C had the highest yield. The optimization results are given in Table 1. The structures of the 16 derivatives of 1,4-dihydropyrano[2,3-c]pyrazole-5-carbonitrile (5a-o) synthesized in this study under optimal conditions are given in Table 2.
As a result, the zirconium magnetic nanocomposite synthesized the desired product with better results, including higher efficiency, a shorter time, and a lower temperature.
As mentioned earlier, the favorable conditions of the zirconium magnetic nanocomposite as a catalyst can be attributed to its high specific surface area.
Another essential advantage of the catalyst studied in this study was its recycling ability. Figure 7 shows that the zirconium magnetic nanocomposite can be reused up to six times (for 5a), which does not significantly reduce the efficiency of the product.

Results of Biological Activity of Zirconium Magnetic Nanocomposite
The high specific surface area of the zirconium magnetic nanocomposite synthesized in this study significantly affected the Gram-positive, Gram-negative, and fungal species. Zirconium magnetic nanocomposite's antimicrobial effects were tested based on the MIC (minimum inhibitory concentration), MBC (minimum bactericidal concentration), and MFC (minimum fungicidal concentration) parameters (Table 4).

Results of Biological Activity of Zirconium Magnetic Nanocomposite
The high specific surface area of the zirconium magnetic nanocomposite synthesized in this study significantly affected the Gram-positive, Gram-negative, and fungal species. Zirconium magnetic nanocomposite's antimicrobial effects were tested based on the MIC (minimum inhibitory concentration), MBC (minimum bactericidal concentration), and MFC (minimum fungicidal concentration) parameters ( Table 4).
The antibacterial effects of nanoparticles were examined on Staphylococcus epidermidis and Bacillus cereus (Gram-positive), Klebsiella pneumonia and Shigella dysenteriae (Gramnegative), and Candida albicans (fungi).
Furthermore, the antimicrobial effects of some commercial drugs (Cefazolin as an antibacterial drug and Terbinafine as an antifungal drug) on the studied species were tested to compare their effectiveness to the zirconium magnetic nanocomposite. The results of the antimicrobial tests proved that the zirconium magnetic nanocomposite positively affected all studied Gram-positive, Gram-negative, and fungi species. The MBC value was 16 µg/mL on Staphylococcus epidermidis, 128 µg/mL on Bacillus cereus, 64 µg/mL on Klebsiella pneumonia, and 128 µg/mL on Shigella dysenteriae, and the MFC value on Candida albicans was 128 µg/mL.
It is noteworthy that Cefazolin was ineffective on Bacillus cereus and Shigella dysenteriae, and Terbinafine was ineffective on Candida albicans. However, the zirconium magnetic nanocomposite had a positive effect.
As mentioned earlier, the unique properties of the zirconium magnetic nanocomposite can be attributed to its high specific surface area, which is the result of its synthesis method.

Conclusions
In the present study, a zirconium magnetic nanocomposite was synthesized using the microwave method. Analyses such as vibrating sample magnetometer curves, X-ray diffraction patterns, scanning electron microscope and transmission electron microscopy images, Fourier transform infrared spectroscopy, and Brunauer-Emmett-Teller N 2 adsorption/desorption isotherms to identify and confirm its structure were performed. The results of the analyses showed that the synthesis method caused uniform morphology and increased the specific surface area of the zirconium magnetic nanocomposite. The synthesized zirconium magnetic nanocomposite was used as a catalyst in the synthesis of 1,4-dihydropyrano[2,3-c]pyrazole-5-carbonitrile derivatives due to its unique properties, including its high specific surface area, which is essential for catalytic applications. The catalytic activity results compared to the previously reported methods for synthesizing 1,4-dihydropyrano[2,3-c]pyrazole-5-carbonitrile derivatives were significant. One of the other advantages of its use as a catalyst is its possible reuse without a noticeable decrease in efficiency. The high specific surface area of the zirconium magnetic nanocomposite resulted in biological activity, which was effective on Gram-positive, Gram-negative, and the studied fungal species. The noteworthy finding of the antibacterial activity was its higher effectiveness compared to the commercially used drugs.

Data Availability Statement:
The authors confirm that the data supporting the findings of this study are available within the article.