Synthesis and characterization of Se doped Fe3O4 nanoparticles for catalytic and biological properties

In this study, Se-doped Fe3O4 with antibacterial properties was synthesized using by a coprecipitation method. The chemistry and morphology of the Se doped Fe3O4 nanocomposite were characterized by energy-dispersive X-ray spectroscopy, field-emission scanning electron microscopy, X-ray diffraction, vibrating sample magnetometry, and Brunauer–Emmett–Teller spectroscopy. The antibacterial activity of the Fe3O4/Se nanocomposite was examined against G+ (Gram-positive) and G− (Gram-negative) bacteria, in the order Staphylococcus aureus, Staphylococcus saprophyticus, Pseudomonas aeruginosa, Klebsiella pneumonia, and Escherichia coli, which are the most harmful and dangerous bacteria. Fe3O4/Se, as a heterogeneous catalyst, was successfully applied to the synthesis of pyrazolopyridine and its derivatives via a one-pot four-component reaction of ethyl acetoacetate, hydrazine hydrate, ammonium acetate, and various aromatic aldehydes. Fe3O4/Se was easily separated from the bacteria-containing solution using a magnet. Its admissible magnetic properties, crystalline structure, antibacterial activity, mild reaction conditions, and green synthesis are specific features that have led to the recommendation of the use of Fe3O4/Se in the water treatment field and medical applications. Direct Se doping of Fe3O4 was successfully realized without additional complicated procedures.

www.nature.com/scientificreports/ at room temperature. The prepared product was collected using an external magnetic field, washed with water and ethanol several times, and dried to afford a brown solid.
Common procedure for the synthesis of pyrazolopyridine derivatives 5a-j. The catalytic activity of the Fe 3 O 4 /Se nanocomposite was tested in a one-pot synthesis of pyrazolopyridine derivatives. The four components were mixed and reacted in the presence of 1 ml EtOH at room temperature: hydrazine hydrate (2 mmol), ethyl acetoacetate (2 mmol), aromatic aldehydes (1 mmol), and ammonium acetate (3 mmol) (Table1). Thin-layer chromatography was used to evaluate reaction completion process (TLC). The undissolved magnetic nanocatalyst was separated from the reaction mixture using a magnet after completion of the reaction. To obtain pure pyrazolopyridine derivatives, the crude product was recrystallized from EtOH. All the products were wellknown chemicals recognized by comparing their melting points with those in the original literature (Table 3).
Spectral data of selected products. Spectral data of selected products. Analysis of the results of NMR. 3, 5-Dimethyl-4-phenyl-1, 4, 7, 8-tetrahydrodipyrazolo [3, 4- Plate-count method. One of the most important and practical methods for determining the concentration of microbes in a sample is to dilute the sample, grow the microbes on the plates and count the colonies. Since the catalyst has been used several times in the reaction in this regard, the primitive pattern of this nanocomposite was compared with the secondary separate pattern of the Fe 3 O 4 /Se nanocomposite after six cycles of use in the organic reaction pyrazolopyridine, as seen in (Fig. S7), XRD analysis of the consumed catalyst underwent changes after six cycles of use, but there are still index peaks related to the crystal structure of the nanocomposite, which confirms the preservation of the catalyst structure. However, new peaks also appeared, which could be due to impurities in the products or reactants on the surface of the recycled catalyst. (See Supplementary Information).
FESEM and TEM analysis. Used to observe particle size distribution, surface morphology, and particle aggregation mode in prepared samples Field-emission scanning-electron microscopy (FESEM) and transmission electron microscopy (TEM) analyses were utilized to examine morphologies and structure. As shown in Fig. 3 4 , the FESEM images of the Fe 3 O 4 /Se NPs are presented at three scales: 1 µm, 500 nm, and 200 nm. The Fe 3 O 4 /Se images showed a spherical structure, in the nanocomposite images, in addition to the spherical structure, the distribution of Se NPs on the Fe 3 O 4 support was also observed. The average particle size of the 35 spherical particles in the nanocomposite was determined to be approximately 80 nm in addition, As is seen in the TEM image, the spots distributed around the magnetic nanoparticles represent the Se, which confirms the good composition of the Se with the Fe 3 O 4 also Se acts as the matrix of the inorganic composite.
The N 2 adsorption-desorption isotherm. The N 2 adsorption-desorption isotherm of Fe 3 O 4 /Se is shown in Fig. 4. Detailed information including the surface area, pore volume, and pore size (width) of the Fe 3 O 4 /Se catalytic system, calculated using the (Brunauer-Emmett-Teller (BET)) and (Barrett-Joyner-Halenda(BJH)) methods is presented in Table 1. The BET surface area and pore volume for Fe 3 O 4 /Se were recorded to be approximately 11.57 m 2 /g and 0.073 cm 3 /g, respectively, which are slightly less than reported values in the literature 66 Antibacterial activity. It was discovered that adding Se NPs to the mix might efficiently boost the generation of reactive oxygen species (ROS) 20 . The most widely accepted mechanism of accomplishment for Se NPs is particle attachment to the bacterial surface and the release of selenium ions into the bacterial cell, which results in oxidative stress, protein synthesis inhibition, or DNA mutation 68 .
There are several putative mechanisms of action of Se NPs. Four possible modes of action were examined to determine whether they explain the antibacterial properties of Se NPs: (1) metabolic intrusion by disturbance All living organisms use adenosine triphosphate (ATP) as an internal energy source. It is the most important energy source for many enzymatic processes; therefore, it is essential for respiration and metabolism. The energyuncoupling effect is characterized by the rapid depletion of cellular ATP. Another mechanism that contributes to bacterial mortality is oxidative stress caused by excessive ROS production in response to nanoparticles 69 .

Catalytic application. Another goal of Fe 3 O 4 /Se nanocatalyst production, as noted in the introduction, is
to investigate its catalytic efficacy in organic processes.
In the synthesis of pyrazolopyridine derivatives, the catalytic activity of Fe 3 O 4 /Se was studied. Different experimental conditions such as temperature, solvent, catalyst amount, and catalyst type were investigated in a one-pot four-component reaction of ethyl acetoacetate (2 mmol), hydrazine hydrate (2 mmol), 2, 4-dichlorobenzaldehyde (1 mmol), and ammonium acetate (3 mmol) as a model reaction to obtain the best result. First, the model reaction was carried out at two different temperatures without the use of a catalyst or solvent, and the yield of the products was 37% ( Table 2, entry 3). The reaction yield was approximately 37% when the Fe 3 O 4 /Se nanocatalyst was added to the model in the absence of a solvent (Table 2, entry 3). Next, ethanol was added to the reaction in the presence of a catalyst at room temperature to analyze the solvent effect, and the efficiency increased significantly ( Table 2, entry 4). The process was then repeated at 80 °C to determine how the temperature affected the outcome, and it was discovered that increasing the temperature to 80 °C caused the reaction to progress (Table 2, entry 5). Subsequently, under reflux and ultrasonic conditions, optimization studies were carried out in H 2 O and EtOH as a green medium; the greatest efficiency was found in EtOH media at room temperature ( Table 2,   , and a wide surface area worked as an efficient catalyst for the one-pot four-component reaction to produce pyrazolopyridine derivatives (Fig. 8).
Examination of the catalytic activity of Fe 3 O 4 /Se. The optimal conditions for synthesizing pyrazolopyridine derivatives using several types of aromatic aldehydes with hydrazine hydrate, ammonium acetate, and ethyl acetoacetate were investigated to assess the generality, application, and limitations between 35 and 90 min at room temperature. All aromatic aldehydes with electron-withdrawing and electron-donating substituents resulted in the synthesis of the corresponding high-yield products, as indicated in Table 3, with no byproducts detected.
Proposed mechanism. The most likely mechanism for the synthesis of different pyrazolopyridine derivatives with Fe 3 O 4 /Se nanoparticles is the four-step mechanism shown in Fig. 9. This is not only due to the abundant Lewis acid sites (Fe 3 + of Fe 3 O 4 ) and, to some extent, the high electrophilic properties of selenium nanoparticles, as well as their physical properties, large surface area, and high thermal and mechanical stability can play an important role in all steps of this four-step reaction illustrated in Fig. 9. Explaining the mechanism of this reaction according to the literature reports 62,70 , first, due to the acidic sites of Fe and the electrophilic property of Se, the oxygen of the carbonyl ethyl acetoacetate groups is involved by the catalyst, which activates the car-       Table S1. The results show that the current technology is superior in terms of catalyst biocompatibility, use of an ecologically friendly solvent, and generation of the desired products with high yields in a reasonable amount of time under mild reaction conditions.

Reusability of Fe 3 O 4 /Se.
In terms of industrial and commercial considerations, catalyst reusability is one of the most significant variables, and the ability to recycle products can largely lead the reaction process to conform to the concepts of green chemistry. The recoverability and reusability of the Fe 3 O 4 /Se nanocatalyst were assessed in the synthesis of 5b. Because of the magnetic nature of the catalyst, it can be easily isolated from the reaction mixture using a magnet bar, repeatedly rinsed with distilled water and ethanol, and then dried after each run. Fortunately, after six consecutive cycles, very little catalyst deactivation occurred during pyrazolopyridine synthesis (Fig. 10). The X-ray pattern of the recycled catalyst was almost identical to that of the fresh catalyst (Fig. S7).

Conclusion
In summary, new, efficient, low-cost, and reusable Fe 3 O 4 /Se NPs were produced and synthesized as nanocomposites using a simple coprecipitation technique. Various analytical methods have been used to evaluate nanocomposite structure, morphology, surface area, pore volume, pore size (width), magnetic properties, and antibacterial characteristics. The SEM images verified the stability and integrity of the spherical structure, as well as the excellent dispersion of Se NPs on the Fe 3 O 4 surface. The magnetic properties of the nanocomposites were confirmed by VSM analysis. Additionally, detailed information, including the surface area, pore volume, and pore size (width) of the Fe 3 O 4 /Se catalytic system, was calculated using the (Brunauer-Emmett-Teller (BET)) and (Barrett-Joyner-Halenda (BJH)) methods. This shows that the synthesized nanocatalyst had a suitable surface area and pore size to promote organic reactions. The nanocomposite was employed as a well-organized and recyclable heterogeneous nanocatalyst for the production of pyrazolopyridine products via a four-component reaction. At room temperature and under mild reaction conditions, high to excellent product yields were achieved using Fe 3 O 4 /Se as a nanocatalyst. The catalyst can be simply removed from the reaction medium by an external magnetic field, washed and dried, and used several times without a substantial loss of activating sites. The antibacterial properties of this nanocomposite were investigated in the removal and destruction of G + S. aureus, S. saprophyticus, G − E. coli, K. pneumonia, P. aeruginosa bacteria, a group of dangerous bacteria that threaten the health of living organisms. This nanocomposite can also be utilized to disinfect water polluted with bacteria; www.nature.com/scientificreports/ the inactivation of S. aureus and E. coli in the presence of nanoparticles was confirmed by the colony method. Moreover, this is the first report on the plan, production, functionalization, and characterization of the current nanocomposite, as well as its presentation as a heterogeneous nanocatalyst in a significant organic process. Pyrazolopyridine is one of the most important heterocyclic compounds with many biological activities. In this paper, we report an effective and practical method for the synthesis of pyrazolopyridine and its derivatives.

Data availability
All data generated or analysed during this study are included in this published article [and its supplementary information files].