Asparagine-EDTA MNPs: A Highly Ecient And Recyclable Magnetic Multifunctional Core-Shell Nanocatalyst For Green Synthesis of Biologically-Active 3,4-Dihydropyrimidin-2(1H)-One Compounds

In this study, the new asparagine grafted on the EDTA-modied Fe 3 O 4 @SiO 2 core-shell (Fe 3 O 4 @SiO 2 -APTS-EDTA-asparagine) magnetic nanoparticles were prepared and their structures were properly conrmed using different spectroscopic, microscopic and magnetic methods or techniques such as FT-IR, EDX, XRD, FESEM, TEM, TGA and VSM. The Fe 3 O 4 @SiO 2 -APTS-EDTA-asparagine core-shell nanomaterial was examined, as a highly ecient multifunctional and recoverable nanocatalyst, for the synthesis of a wide range of nitrogen-containing heterocycles and biologically-active 3,4-dihydropyrimidin-2(1H)-one derivatives under solvent-free conditions. It was proved that Fe 3 O 4 @SiO 2 -APTS-EDTA-asparagine MNPs, as a catalyst having excellent thermally and magnetic stability, specic morphology and acidic sites, can activate the Biginelli reaction components. Moreover, environmental-friendliness and nontoxic nature properties of the catalyst, cost effectiveness, low catalyst loading, easy separation of the catalyst from products and short time of reaction are some of the remarkable advantages of this green protocol.

in ammatory, antihypertensive agents, calcium channel blockers, antitumor compounds [41][42][43][44][45][46][47] . A simple and general protocol for access to 3,4-dihydropyrimidin-2(1H)-ones involves a three-component one-pot Biginelli cyclocondensation of ethyl acetoacetate, urea and various aldehydes accelerated by different types of catalytic systems such as polymer-supported catalysts 48 , ionic liquids 49,50 , ionic liquid/silica sulfuric acid 51 , metal−organic framework (MOF) 52,53 , montmorillonite clay 54 , magnetic nanoparticles 55 , Lewis acidic zirconium (IV)-salophen per uorooctanesulfonate or sulfated polyborate 56,57 , nanocrystalline CdS thin lm 46 , graphene oxide 58,59 and mesoporous materials 60,61 as well as environmental friendly energy inputs such as ultrasound 62 or microwave irradiation 63 . Most of the reported methods in this regard have the role of heterogeneous catalysts and high value. However, these have problems such as complicated and tedious separation of products and catalysts, toxic reaction conditions, long reaction times and low yields. Therefore, there is still room to develop more environmentally-benign protocols to promote the Biginelli MCR condensation.
In many previous reports, ethylenediaminetetraacetic acid (EDTA) has been used as an ion exchange and chelating agent for various metal ions [64][65][66] , but this compound has a good ability as an inexpensive and non-toxic cross-linker to make strong bonds with organic materials having nucleophilic centers 67 . On the other hand, asparagine is one of the 20 amino acids found in the cells of the human body and is essential for maintaining balance in the central nervous system 68 . Asparagine can act as a biocompatible precursor and bifunctional organocatalyst due to its high natural abundance and costeffectiveness with acidic and basic sites 69,70 .
In this research, we herein report the synthesis and characterizations of new asparagine grafted on the EDTA-modi ed Fe 3 O 4 @SiO 2 core-shell magnetic nanoparticles (Fe 3 O 4 @SiO 2 -APTS-EDTA-asparagine), as a magnetically recoverable nanocatalyst, to promote the Biginelli reaction e ciently at 60°C under solvent-free conditions (Fig. 1).
The FT-IR spectroscopy was employed to determine the functional groups and structure of Fe 3 O 4 (a), Fe 3 O 4 @SiO 2 (b), Fe 3 O 4 @SiO 2 -APTS (c), Fe 3 O 4 @SiO 2 -APTS-EDTA (d) and Fe 3 O 4 @SiO 2 -APTS-EDTA-asparagine (e). The results are presented in Fig. 2. In the spectra of Fe 3 O 4 nanoparticles (Fig. 2a) the bands displayed at 620 cm −1 and about 3410 cm −1 are attributed to stretching vibration of Fe−O bond and surface hydroxyl groups, respectively. These peaks were observed in all ve samples isolated at the different synthetic stages. In the FT-IR spectrum of Fe 3 O 4 @SiO 2 (Fig. 2b), the absorption bands at 881 and 1036 cm −1 can be ascribed to the presence of Si−O−Si symmetric and Si−O−Si asymmetric stretching modes, re ecting the coating of silica layer on the magnetite nanoparticles 71 . SP 3 C-H stretching vibrations about 2922 cm −1 con rmed the presence of the anchored (3-aminopropyl) triethoxysilane (APTS) group and the band about 1400 cm −1 is assigned to the bending of −NH groups of Fe 3 O 4 @SiO 2 -APTS MNPs (Fig. 2c) 72 . In the FT-IR spectrum of Fe 3 O 4 @SiO 2 -APTS-EDTA (Fig. 2d), the peaks at 1635 cm −1 , 1707 cm −1 and 1760 cm −1 corresponding to the C=O vibration of amide, acid and anhydride groups, respectively. In the last step, the peak at 1760 cm −1 , which belongs to the anhydride group has been removed and new peaks at 1651 cm −1 and 1737 cm −1 are attributed to the amide and acid groups in the surface of Fe 3 O 4 @SiO 2 -APTS-EDTA-asparagine (Fig. 2e). These results from the FT-IR spectrum con rm that the silica coating and subsequent steps have been successfully performed on the surface of Fe 3 O 4 .
The morphology and texture of Fe 3 O 4 @SiO 2 -APTS-EDTA-asparagine MNPs (1) were indicated by FESEM analysis and their photographs were presented in Fig. 4. According to these FESEM photographs, the size and surface shape of nanoparticles are well observed, which proves that the particles are spherical and without agglomeration. The FESEM photographs supported the formation of spherically shaped MNPs, which is in accordance with TEM analysis.
The TEM analysis of the Fe 3 O 4 @SiO 2 -APTS-EDTA-asparagine (1) MNPs in two scales is shown in Fig. 5.
The TEM images demonstrated structural order and the morphology suggested that the magnetite nanoparticles have an average diameter size of 41 nm.
The XRD pattern of Fe 3 O 4 @SiO 2 -APTS-EDTA-asparagine (1) was shown in Fig. 6. The re ection peaks were compared with the reference standard patterns related to EDTA (card no. JCPDS, 00-033-1672),  (1) is su cient to be recovered by exerting an external magnet.
Thermal stability of the Fe 3 O 4 @SiO 2 -APTS-EDTA-asparagine nanomaterial (1) was investigated under the air atmosphere over the temperature range of 50 − 800°C (Fig. 8). The Fe 3 O 4 @SiO 2 -APTS-EDTAasparagine MNPs (1) display three weight loss steps over the temperature range of TGA and the total weight loss of nanocatalyst 1 is around 60%. According to obtained results, in the rst step 15% weight loss in the range of 150 − 200°C is due to the evaporation of adsorbed water and organic solvents that remain in the nanocatalyst through its preparation processes. In addition, 22% weight loss in the range of 200 − 400°C corresponds to the loss of EDTA-asparagine moiety. In the last step, the sharp weight loss of 23% at 400-700°C can be assigned to the decomposition of APTS moiety in the MNPs framework. These results also indicate that APTS, EDTA and asparagine has been successfully grafted onto the surface of Fe 3 O 4 @SiO 2 . Above 700°C only Fe 3 O 4 was present.
Optimization of conditions in the Biginelli reaction using Fe 3 O 4 @SiO 2 -APTS-EDTA asparagine nanocatalyst (1) In our preliminary experiments, the catalytic activity of as prepared catalyst 1 was evaluated in the formation of dihydropyrimidin-2(1H)-one derivatives by the Biginelli condensation. For this purpose, reaction conditions were optimized using the equimolar mixtures of urea (2, 1 mmol), 4chlorobenzaldehyde (3a, 1 mmol) and ethyl acetoacetate (4a, 1 mmol) as the model reaction (Eq. 1). In a systematic screening, the reaction conditions were investigated precisely by considering of several crucial variables such as catalyst loading, reaction time, solvent and reaction temperature, as given in Table 1.
Initially, in the absence of any catalyst and solvent, the progress of model reaction was slow and the yield of the 9-(4-chlorophenyl)-3,3,6,6-tetramethyl-3,4,6,7,9,10-hexahydroacridine-1,8(2H,5H)-dione (5a) was trace, even after a long time ( Table 1, entry 1). Then, in the presence of very low amount of Fe 3 O 4 @SiO 2 -APTS-EDTA-asparagine (1) loading, as a nanocatalyst, without any solvent at room temperature, a good yield of the desired product 5a was obtained ( Table 1, entry 2). To achieve an excellent yield, the reaction temperature was increased to 60°C (Table 1, entry 3). Afterward, the model reaction was performed with lower catalyst 1 loading under solvent-free conditions as well as polar and non-polar solvents. Furthermore, the effect of temperature and different solvents was investigated (Table 1, entries 4-13). Also, the model reactions in the presence of EDTA and asparagine were separately investigated, but lower yields of the desired product 5a were isolated ( Table 1, entries 14-15).
Following the steps of optimizing the reaction conditions, the effect of different solvents and amount of catalyst loadings are summarized in Fig. 9. The model reaction was investigated under solvent-free conditions and different solvents such as EtOH, MeOH, EtOH/H 2 O (1:1), and DMF using Fe 3 O 4 @SiO 2 -APTS-EDTA-asparagine nanocatalyst (1) with different loading of the catalyst 1. According to the obtained ndings summarized in Table 1 and Fig. 9, the optimum reaction conditions were found to be 10 mg of Fe 3 O 4 @SiO 2 -APTS-EDTA-asparagine nanocatalyst (1) loading under solvent-free conditions at 60°C . After the above experiments, the scope of reaction was expanded by using aromatic aldehydes having electron-withdrawing or electron-donating groups under the optimized conditions (Eq. 2). The results are summarized in Table 2. As expected, in this novel magnetic heterogeneous catalytic system the reaction rate of aldehydes with electron-donating groups was slower than electron-releasing ones and required more time to complete the reaction. An alternative variation in this reaction was accomplished by utilizing methyl acetoacetate (4b) instead of ethyl acetoacetate (4a) for the synthesis of different Biginelli products. It is worth noting that all the reactions represented very good to excellent yields under solventfree conditions in short time.
The proposed mechanism for the synthesis of 3,4dihydropyrimidin-2(1H)-one derivatives in the presence of

Green chemistry metrics
In this part of our research, green chemistry metrics for the synthesis of 3,4-dihydropyrimidin-2(1H)-one by Fe 3 O 4 @SiO 2 -APTS-EDTA-asparagine nanocatalyst (1) were calculated and the results are summarized in Table 3 79,80 . Hence, several parameters of the green approach such as environmental factor (E factor), process mass intensity, reaction mass e ciency, carbon e ciency, and atom economy were evaluated and compared to the ideal values 81 . As presented in Table 3, all calculated values are close to the ideal values and were reported in supporting information.
Reusability of the Fe 3 O 4 @SiO 2 -APTS-EDTA-asparagine nanocatalyst (1) One of the critical scales in catalytic processes is reusability and recyclability of the catalyst. For evaluation of this parameter, the model reaction was examined using Fe 3 O 4 @SiO 2 -APTS-EDTAasparagine (1) for four runs. At the end of the reaction, the catalyst 1 was removed using an external magnet and the recycled catalyst was washed with dry toluene, dried and used in a subsequent model reaction. The obtained results are summarized in Fig. 11. Considering the results of isolated yields of products, the catalytic activity of Fe 3 O 4 @SiO 2 -APTS-EDTA-asparagine nanocatalyst (1) after four runs is slightly reduced, which demonstrates proper conservancy of the catalytic activity after recycling.

Comparative study of Fe 3 O 4 @SiO 2 -APTS-EDTA-asparagine nanocatalyst (1) and other catalysts for the Biginelli reaction
In order to compare the optimal catalytic activity and reaction conditions of the Fe 3 O 4 @SiO 2 -APTS-EDTAasparagine nanocatalyst (1) with previously reported catalysts for the three-component Biginelli reaction, we compared reaction conditions and yield of desired product (5a) in Table 4. As it can be observed from data in Table 4, all catalytic systems are capable of producing the desired product with satisfactory yields but Fe 3 O 4 @SiO 2 -APTS-EDTA-asparagine nanocatalyst (1) in terms of yield and time factors, the reaction temperature, solvent and amount of catalyst loading demonstrates better performance than the other catalysts. Furthermore, additional advantage of this protocol is its easy separation from the crude products by using an external magnet compared to the most of reported heterogeneous catalytic systems.  (1) were recorded by Bahr company STA 504. X-ray diffraction (XRD) pattern of the catalyst 1 was taken by the Bruker D8 Advance device. The composition of the catalyst was determined by energy-dispersive Xray (EDX) spectroscopy using a Numerix DXP-X10P instrument. Magnetization measurements were carried out on a BHV-55 vibrating sample magnetometer (VSM). 1 H NMR spectra of the isolated products were recorded at 500 MHz using a Varian-INOVA spectrometer.

Chemicals and Instrumentation
General procedure for preparation of the magnetic Fe 3 O 4 nanoparticles Preparation of Fe 3 O 4 nanoparticles were according to a reported general method 83 . In this procedure, in a 100 mL round-bottomed ask FeCl 3 .6H 2 O (4.6 g, 0.017 mol) and FeCl 2 .4H 2 O (2.3 g, 0.011 mol) were dissolved in deionized water (60 mL) and stirred for 30 min. Subsequently, aqueous NH 3 (10 mL) was added dropwise into the mixture and heated to 40°C under N 2 atmosphere for 2 h. The black solution was poured from the reaction vessel and Fe 3 O 4 MNPs precipitates were separated from the solution using an external magnet, washed ve times with deionized water and EtOH, and dried in the oven at 50°C for 24 h.
General procedure for preparation of the silica-coated magnetic nanoparticles (Fe 3 O 4 @SiO 2 ) In accordance to the modi ed Stöber method, silica-coated Fe 3 O 4 nanoparticles (Fe 3 O 4 @SiO 2 ) were produced by a solvothermal reaction 84 Table 2. After completion of the reaction, as monitored by TLC [eluent: nhexane: EtOAc: 3:1], the catalyst was separated using an external magnet and the residue was concentrated to result in the crude product. Finally, the crude product was recrystallized from EtOH to obtain the pure product.

Conclusion
In summary, the novel and thermally stable asparagine grafted on EDTA-modi ed Fe 3 O 4 @SiO 2 core-shell magnetic nanoparticles (Fe 3 O 4 @SiO 2 -APTS-EDTA-asparagine) was prepared for the rst time. The Fe 3 O 4 @SiO 2 -APTS-EDTA-asparagine heterogeneouse nanocatalyst was used for highly e cient, facile, and green and sustainable synthesis of 3,4-dihydropyrimidin-2(1H)-one derivatives in a one-pot and threecomponent protocol through cyclocondensation of alkyl acetoacetate, urea and various aldehydes under solvent-free conditions. Consistency with the ideal values of green chemistry parameters, easy work up procedure, good to excellent yields in shorter reaction times, fast separation and recyclability of the catalyst are the additional advantages for its application in academic and industrial purposes. class of carbazolyl dihydropyrimidinones via an improved Biginelli protocol. New Journal of Chemistry 43, 10989-11002 (2019    TGA curve of the magnetic Fe3O4@SiO2-APTS-EDTA-asparagine nanomaterial (1).

Figure 9
Effect of solvent and the amount of Fe3O4@SiO2-APTS-EDTA-asparagine nanocatalyst (1) on the model reaction.