Synthesis of a novel ternary ZIF-8/GO/MgFe2O4 nanocomposite and its application in drug delivery

In recent year, metal–organic frameworks (MOFs) have been displayed to be a category of promising drug delivery systems because of their crystalline structure, the potential of further functionality, and high porosity. In this research, graphene oxide was synthesized from pure graphite via hummer method and then MgFe2O4 nanoparticles was incorporated into the synthesized ZIF-8 metal–organic frameworks which followed with loading on the surfaces of graphene oxide. In continue, tetracycline as an antibiotic drug was loaded on the surfaces and the cavities of the prepared nanocomposite. The outcomes of this research revealed that 90% of the tetracycline was loaded on the synthesized ZIF-8/GO/MgFe2O4 nanostructure. Next, drug release was done at pH: 5 and pH: 7.4 within 3 days, resulting about 88% and 92% release of the tetracycline, respectively. With using different spectroscopic methods like X-ray crystallography (XRD), scanning electron microscope (SEM), energy-dispersive X-ray spectroscopy (EDX/Mapping), Fourier transform infrared (FTIR), thermalgravimetric analysis (TGA), and Brunauer–Emmett–Teller (BET), the structure of synthesized materials was confirmed. Furthermore, the antibiotic activity of tetracycline trapped into the ZIF-8/GO/MgFe2O4 was evaluated by agar-well diffusion method on both gram-positive (Staphylococcus aureus) and gram-negative (Escherichia coli) bacteria, which showed good antibacterial results.

In last decade, more attention has been paid to improvement novel techniques for the advancing of drug delivery systems. The increasing numbers of drug resistances in bacterial contaminants have become important healthcare challenges due to the severe reduction in the number of therapy and repetitive treatment accessible. This issue has led to enhanced morbidity developed medicinal. As a result, combination treatment involving the co-application of existing antibiotics with unique nano structures such as metal halides, nano-sized transporters, and metal oxide nanoparticles is a promising approach to counter antimicrobial opposition 1 .
The tetracycline (TC) is a type of antibiotics that constrain protein combination by stopping the addition of aminoacyl-tRNA to the ribosomal acceptor (A) sites ( Fig. 1) 2 .
TCs are broad-spectrum agents, displaying activity against a varied kind of bacteria 3 . The favorable antimicrobial and antifungal attributes of these agents and the loss of mighty adverse side effects has led to their applications in the treatment of animal and human infections. The main feature of this drug is its use at certain hours in order to completely control the infection 2 .
Nano-scale carbon allotrope have been extensively investigated by researchers for targeted drug delivery. In this regard, graphene sheets with highly active surfaces are astonishingly desirable for fundamental research and technological applications. This two-dimensional structure has been considered due to having several functional groups connected to carbon plates with high density, biocompatibility, high electrical, and conductivity properties 4 .
Graphene oxide (GO) as a kind of graphite derivatives give easy dispensability in water and other organic solvents. GO possess wealthy oxygen containing functionalities such as epoxide, carboxyl, ketone, lactone, and hydroxyl groups 5 . Furthermore, it has a great specific surface area and displays excellent activities which are a versatile tool in chemical transformations 6 , and also in drug delivery systems 7 .
Metal-organic frameworks have lately concerned very attention in the meantime. MOFs display many basic specifications in drug delivery due to flexible building, variable sizes and figures, easily functionalization, high porosity, easy biodegradability and high design capability [8][9][10] . ZIF-8 is a type of zeolitic imidazole frameworks (ZIFs) which have excellent benefits because of highly pore sizes, massive surface area and simple variation of www.nature.com/scientificreports/ the pore sizes inside the frameworks with modifiable organic groups 11 . However, to improve the stability of the carrier, scientists have been used some metals with high oxidation state (Ni 2+ , Al 3+ , Zr 4+ , Fe 3+ , Mg 2+ ) to form powerful coordination bond in ligands to provide extremely stable MOFs. Therefore, great oxidation state metalbased MOFs or their composites display considerable potential in antibiotic sensing 12,13 . We used MgFe 2 O 4 nanoparticles with crystal structure in which the particles are rather disordered than crystalline particles, there by having many more defects can provide better nanocarrier concentration and extra active sites. Also, magnetic nanoparticle MgFe 2 O 4 exhibited greater super paramagnetic propriety and homogeneous diameter. This information indicates that Mg (II) is safe and practical to use in a drug delivery systems 14 .
Previously, metal-organic framework skeletons have been used as potential nanocarriers in drug delivery of some antibiotics 1,15 , folic acid 16 , anticancer drugs 17,18 and ibuprofen 19 .
Considering the significance of the previous issues associated to discovery of new drug delivery systems to find a suitable and practical nanocarrier 20-22 , we were capable to introduce a unique and impressive nanostructure like ZIF-8/GO/MgFe 2 O 4 as a carrier for the loading and release of TC.
Furthermore, the antibacterial activities of the prepared composites were compared and evaluated with pure tetracycline against Staphylococcus aureus and Escherichia coli bacteria by agar-well diffusion method which exhibited considerable results. The target of the this work was to progress the performance of the antibacterial drug delivery method, and drug release control (Fig. 2 (10 mL). In another vessel, 2-methyl imidazole (1.3 g) was poured in methanol (10 mL) and then the mixture was stirred vigorously. Then, two vessels were mixed together and the solution was mixed completely for 30 min. The obtained residue was separated by centrifugation method and was washed several times with methanol. Eventually, the product was dried at 100 °C for 24 h 24 .
Synthesis of graphene oxide. GO was prepared by Hummer's method. Normally, pure graphite (5 g) and sodium nitrate (2.5 g) was added to sulfuric acid (115 mL, 98%) and the solution was equipped to a magnetic stirrer with a condenser which placed in an ice bath. During the stirring of the mixture, KMnO 4 (15 g) was added slowly for 120 min. The mixture was placed in a water bath (35 °C) and stirred for half hour. Next, 230 mL of ultrapure water was gradually added into the mixture and stirring was continued at 98 °C for 15 min. In continue, 700 mL of DI water and H 2 O 2 (50 mL, 30%) were respectively added to the solution to end the reaction. When the reaction completed, the product was washed with HCl (5%) and DI water for three times. The obtained GO was dried at 60 °C for 12 h 25 .  Field-emission scanning electron microscope analysis. The particle size and morphology of the ZIF-8/GO/MgFe 2 O 4 before and after TC loading are indicated in Fig. 4. SEM illustration of the ZIF-8/GO/ MgFe 2 O 4 shows that nanoparticles have almost spherical structure (Fig. 4a). In Fig. 4b, with loading of the TC on the surfaces of the ZIF-8/GO/MgFe 2 O 4 nanocomposite, the particle size was increased and also the morphology of the nanocomposite disordered due to drug loading.

Preparation of ZIF
Energy-dispersive X-ray spectroscopy. EDX technique was applied to explore the elemental components of the prepared ZIF-8/GO/MgFe 2 O 4 /TC nanocomposite. Figure 5 shows that the elements are including Mg, O, Fe, C, N, and Zn for the ZIF-8/GO/MgFe 2 O 4 /TC nanostructure.
Furthermore, EDX analysis has been performed in the mode of elemental mapping of the ZIF-8/GO/ MgFe 2 O 4 /TC nanocomposite (Fig. 6). The highly dispersive of elements distribution verified that there is no impurity in the prepared nanocomposite. It was found that in the item of C (Fig. 6a), Fe (Fig. 6b), N (Fig. 6c), O (Fig. 6d), Zn (Fig. 6e), and Mg (Fig. 6f) which not only exhibited the excellent purity but also disclosed the homogeneous scattering of elements within the ZIF-8/GO/MgFe 2 O 4 /TC. Eventually, Fig. 6g shows the homogeneous arrangement of the elements throughout the structure. Also, SEM imaging in Fig. 6h (10 nm) revealed a few micrometer-sized precipitates, which could not be seen in the SEM images (Fig. 4) due to lower sampling depth.

Brunauer-Emmett-Teller techniques.
The BET absorption technique is a suitable method to calculate the surface area and porosity of the structures. As shown in the BET plots (Fig. 7), the available surface areas were 504.56 m 2 g −1 (Fig. 7a) and 131.06 m 2 g −1 (Fig. 7b) before and after loading TC, respectively. The pore capacity of the cavities in the ZIF-8/GO/MgFe 2 O 4 is 0.3378 cm 3 g −1 which has been decreased to 0.059053 cm 3 g −1 after encapsulation of the TC. This information confirm that the encapsulation of the TC in the cages of ZIF-8/ GO/MgFe 2 O 4 .
Also, BET analysis was applied to investigate the adsorption/desorption of ZIF-8/GO/MgFe 2 O 4 before and after loading with by TC. As displayed in Fig. 8, the adsorption-desorption isotherm of ZIF-8/GO/MgFe 2 O 4 is  www.nature.com/scientificreports/ type I 28 . The results of the BJH technique indicate that the average pore diameter are 1.21 nm, before and after drug loading (Fig. 9).
X-ray diffraction analysis. XRD technique was used to verify the structure of the prepared nanoparticles. . In XRD pattern of ZIF-8 the peaks at 7.5°, 12.5°, and 18° (2θ°) show the formation ZIF-8 metal-organic frameworks (Fig. 10b). The XRD analysis associated with ZIF-8/MgFe 2 O 4 showed that both of the synthesized materials were crystalline and their shapes is consistently based on single crystal techniques (Fig. 10c). The study of the XRD pattern of the ZIF-8/GO/MgFe 2 O 4 (Fig. 10d), revealed that the presented peaks at 9.3° and 11.3° (2θ°) indicated the satisfactory cordination of the GO in the ZIF-8/MgFe 2 O 4 composite. The XRD pattern of the final structure including ZIF-8/MgFe 2 O 4 /GO/TC (Fig. 10e), conforms the peaks at 30.1°, 35.2°, 43.1°, 57.0°, and 62.6° (2θ°) indicate the formation of the MgFe 2 O 4 NPs. Also, the presence peaks at 7.5°, 12.5°, and 18° (2θ°) show the formation of the ZIF-8 frameworks, as well as, the peaks at 9.3° and 11.3° (2θ°) indicate the construction of the GO 30 . From this information it can be concluded that after loading by TC, the crystallite structure of ZIF-8/MgFe 2 O 4 / GO is preserved.    (Fig. 11). The weight loss about 8%, beginning from 220 to 400 °C is recognized to the devastation of TC. Moreover, the breakdown of the MOF was detected at the range of 400-500 °C. Eventually, destruction up to 800 °C is attributed to the GO 31 .
Study on tetracycline release. To calculate the amount of TC loading on the nanocarrier, various concentrations of the TC were made in two buffers (pH: 5 and pH: 7.4) solutions and the absorptions were got by a UV-Vis device at a wavelength of 360 nm. Next, their calibration curves were plotted. The release diagrams at pH: 5 and pH: 7.4 are shown in Fig. 12. As displayed, the stability of ZIF-8/GO/MgFe 2 O 4 /TC nanocarrier at pH: 5, which was quickly destroyed in comparison with pH: 7.4, probably due to the destruction of the structure the TC release at pH: 5 was very quicker than pH: 7.4. The achieved consequences from Fig. 12 display that the percentage of the TC release at pH: 5 increased to 80% in the first 6 h. Although, the release of the TC at physiological of the body (pH: 7.4) was near 76% over 40 h.
The results of the antimicrobial tests. Antimicrobial activities of the tetracycline and the prepared structures were performed on standard strains such as of S. aureus and E. coli. As indicated in Fig. 13 and Table 1, among various structures such as pure tetracycline, ZIF-8/GO/MgFe 2 O 4 , and ZIF-8/GO/MgFe 2 O 4 /TC, the superlative outcomes were got in the presence of ZIF-8/GO/MgFe 2 O 4 /TC as a strong antibacterial agent. In this research, the diameter of the zone of inhibition was evaluated after incubation to conclude the antimicrobial activity. The results revealed that TC loaded on the MOF was as well as inhibited the development of both   Table 1, inhibition zone of 22 mm against E. coli and 25 mm related to S. aureus, such results are obtained when the tetracycline of an inhibitory behavior is shown against S. aureus and E. coli. With an inhibition zone of 17 mm and 18 mm, respectively.

Conclusion
The outcomes of this study revealed that the novel nanostructure including ZIF-8/GO/MgFe 2 O 4 was successfully synthesized based on the characterization and structure elucidation by SEM, EDX/Mapping, XRD, BET, FT-IR, and TGA analysis. In addition, the tetracycline as an antibiotic drug was encapsulated into the ZIF-8/ GO/MgFe 2 O 4 with high loading of 90%, due to the porous nanocomposite, great surface area and cavities in the structure of nanocarrier. In addition, the TC release from the nanocomposite were 88% and 92%, phosphate buffer saline and acetate buffer, respectively. In order to evaluate the antibacterial activities of the prepared composites toward pure tetracycline, the results of by agar well diffusion showed that the antibacterial activity of ZIF-8/GO/MgFe 2 O 4 /TC is more than ZIF-8/GO/MgFe 2 O 4 and TC.