A New Supported Manganese-Based Coordination Complex as a Nano-Catalyst for the Synthesis of Indazolophthalazinetriones and Investigation of Its Antibacterial Activity

A new magnetic supported manganese-based coordination complex (Fe 3 O 4 @SiO 2 @CPTMS@MBOL@ Mn) was prepared in consecutive stages, characterized via various techniques (VSM, SEM, TEM, XRD, FT-IR, EDX, TG-DTA and ICP) and used as a capable nano-catalyst for the fast and green synthesis of divers 2H-indazolo[2,1-b]phthalazine-1,6,11(13H)-triones in a simple and ecient procedure from the one pot three-component condensation reaction of aldehydes, dimedone and phthalhydrazide in reuxed ethanol in excellent yields and fast reaction times. The Mn catalyst can be recycled and reused without notable loss of the catalytic activity. Also, the antibacterial properties of the Mn supported nano-catalyst were studied against a number of gram-positive and gram-negative bacterial strains.


Introduction
Phthalazine derivatives have an important role in heterocyclic compounds that have received terri c interests in the eld of pharmaceutical and biological activities that leads to clinical application 1 such as antifungal 2 , anti-in ammatory, 3 anticonvulsant, 4 anticancer, 5 cardiotonic. 6 In addition, these compounds represents speci c optical properties. 7 Multi-component reactions (MCRs) are notable mechanism for preparation of wide variety of organic compounds via better e ciency in one step with at least three or more starting materials. 8 Using a large number of different type of catalysts which accelerate or proceed the reaction in smooth condition have been extensively developed for these reactions. [9][10][11] Several synthetic routes have been reported for the synthesis of Phthalazine derivatives such as one-pot three-or four-component condensation reactions. [12][13][14][15][16] For example, Fe 3 O 4 @Cys-SO 3 H was used by Kefayati and coworker as a catalyst for the one-pot three-component condensation of aromatic aldehydes, phthalhydrazide and 1,3-dicarbonyl compounds for the synthesis of 2H-indazolo [2,1-b]phthalazinetriones derivatives in ethanol at re uxed condition. 17 Shirini and coworkers applied [PVPH]ClO 4 as an e cient and reusable solid acid polymeric catalyst for the synthesis of 2H-indazolo [2,1-b]phthalazine-triones in the same way of using initial regents through the one-pot reaction at 100°C in solvent-free conditions. 18 Heirati and coworkers investigated implementation of O-sulfonic acid poly(4-vinylpyrrolidonium) chloride as an effective catalyst for the same synthesis at 80°C in solvent-free conditions. 19 Rostamnia and co-worker synthesized a series of 1H-indazolo [1,2-b]phthalazine-1,6,11-triones by similar regents and using the Fe 3 O 4 @GO-Pr-SO 3 H catalyst in re uxed ethanol. 20 5-sulphosalicylic acid was used by Karhale and coworkers as an organocatalyst catalyst for the one-pot synthesis of 2H-indazolo[2,1b]phthalazinetriones from the previous substrates. 21 Varghese and coworkers through a noticeable method synthesized 2H-indazolo[2,1-b]phthalazine-1,6, 11(13H)-triones by similar regents in the presence of iodine under ultrasonic irradiation. 22 Mosaddegh and coworkers succeeded to synthesize phthalazinetriones derivatives through a rapid, one-pot, four-component route by using Ce(SO 4 ) 2 · 4H 2 O at 125°C and under solvent free condition. 23 However, some methods suffer from some limitations such as tedious workup procedures, using corrosive catalysts, low product yields, long reaction times, formation of side products and di culties in recovery of catalysts. 24 Due to concerns on environmental issues different strategies were applied to reduce side-effects of chemical industries, for instance using solvent-free techniques, eco-friendly materials, ultrasound or microwave methods and nally recyclable catalysts. 25 In recent years the application of magnetic nanoparticles (MNPs) according to the super magnetic property and facile separation from reactions media, high surface area, low toxicity, biocompatibility, good stability, low cost has obtained noticeable regards for chemists using as catalysts in chemical reactions. Moreover, MNPs can be functionalized completely via proper surface modi cations. Based on mentioned properties, many MNPs-supported catalysts have been effectively applied for improving a large number of chemical reactions. 26, 27 Among the heterogeneous catalysts, the magnetic supported metal complexes have special places, 28 so we would like to announce the synthesis a novel Mn supported nano-catalyst  to 1602 cm − 1 which could be in accordance of the new interaction of manganese with the nitrogen of the C = N bond. Also at 408.93 cm − 1 a weak peak was appeared in corresponding to the Mn-N band. Therefore, comparison of the all IR spectra approved the successful formation of the Mn supported nanocatalyst.

EDX analysis of the Mn supported nano-catalyst
The elemental structure of the Mn supported nano-catalyst was disclosed by the EDX analysis (Fig. 3).
The results rati ed the existence of the predictable elements in the construction of the catalyst, including C, N, O, Si, S, Fe and Mn. Generally the sample will be usually coated with a very thin layer of gold that leads appearing of the Au peak in the EDX graph analysis, as well.

SEM and TEM analysis of the Mn supported nanocatalyst
In another evaluation, the morphology and size of the Mn supported nano-catalyst were determined by SEM and TEM images ( Fig. 4-5). The spherical shape of the Fe 3 O 4 @SiO 2 @CPTMS@MBOL@Mn is conceived through the uniform nanometer-sized particles. In the SEM images, formation of sintered grains with the 35-47 nm size range was apparent. The core-shell structure of nano catalyst could be observed via TEM images. However, magneto static interactions of the particles could be the reason of some particle aggregations. At an exact scrutiny, according to distribution histograms (Fig. 6), the average sizes of the nanoparticles is anticipated between 19.1 and 29.7 nm.

Thermal properties of the Mn supported nano-catalyst
The thermo-gravimetric analysis shows the thermal stability and decomposition behavior of the Mn supported nano-catalyst beyond heating (Fig. 6). As observed there are about four weight loss stages for the catalyst in the ranges of 100, 185, 300, and 360 C, respectively. The initial weight reduction at about 100 C probably corresponds to the residual water, the second weight loss at about 180 C is possibly attributed to the thermal decomposition of the complex, the third weight loss at about 300 C is attributed to the thermal decomposition of the two CPTMS and MBOL ligands, and the fourth weight reduction at about 360 C is probably resulted from the de-coating of Fe 3 O 4 .

VSM analysis of the Mn supported nano-catalyst
Due to comparison of magnetic properties, the VSM analysis was performed for the ve steps of preparing including A) Fe 3 O 4 MNPs, B) Fe 3 O 4 @SiO 2 , C) Fe 3 O 4 @ SiO 2 -CPTMS, D) Fe 3 O 4 @SiO 2 @CPTMS@MBOL and E) Fe 3 O 4 @SiO 2 @CPTMS@MBOL@Mn (Fig. 7). Generally, all ve steps of formation show magnetic possessions and a clear decrease from A to E (65, 35, 30, 10 and 4.35 emu/g, respectively) can be observed. In fact the dipolar-dipolar interactions reduce between the magnetic nanoparticles due to coating of Fe 3 O 4 MNPs via different layers and complexation.

General
All chemicals (starting materials, reagents, solvents, etc) were acquired from the chemical companies and used in the absence of additionally puri cation.

Preparation of the Mn supported nano-catalyst
The Mn supported nano-catalyst was prepared according to the procedure described. 32 3.3 General Procedure for the synthesis of indazolophthalazinetriones Phthalhydrazide (0.162 g, 1.0 mmole), dimedone (0.140 g, 1.0 mmole), an aldehyde derivative (1.0 mmole) and the Mn supported nano-catalyst (20 mg) were mixed in ethanol (5 mL), stirred and heated at re ux condition in appropriate times which depends on the substrate (Table 2), and completion of the reaction was monitored with TLC. After cooling, the Mn supported nano-catalyst was separated by a super magnet, H 2 O (10 mL) added, and a white precipitate ltered and recrystallized in EtOH.

Optimization
To obtain the best result, effects of different conditions were investigated for this reaction.  Also, two additional reactions were performed under the optimum conditions as below: 1. The synthesis of indazolophthalazinetriones carried out in the absence of Fe 3 O 4 @SiO 2 @CPTMS@MBOL@Mn: The reaction e ciency was very low (trace).
2. The reaction was done in the presence of (Fe 3 O 4 @SiO 2 @CPTMS@MBOL) and no product was observed.

Synthesis of diverse indazolophthalazinetriones (2a-j)
According to the optimization of model reaction, different indazolophthalazinetriones (2a-j) were synthesized from the one pot three-component condensation reaction of aldehydes, dimedone and phthalhydrazide in EtOH at re ux conditions in attendance of the Mn supported nano-catalyst with good to excellent yields in fast reaction times (Table 2).

Characterization of the products
All indazolophthalazinetriones were characterized and recognized by considering to their physical and spectroscopic analysis and comparing the reported in the literature. The structures of all products were con rmmed via their IR and NMR spectra (Supplementary data).

The plausible mechanism
A persuasive mechanism for the synthesis of diverse phthalazine-triones is illustrated as below. The reaction is followed by mixture of regents in EtOH at re ux conditions in the presence of the Mn supported nano-catalyst (Scheme 3).
The suggested mechanism is presumably in accordance of the Lewis acidity of the Mn supported nanocatalyst by connecting to the carbonyl group to simplify the nucleophilic attack of the enolic form of dimedone (Knoevenagel condensation) to aldehyde to form the intermediate with the subsequent deletion of water, the nucleophilic attack of phthalhydrazide, deletion of water, and the nal cyclization to get the product. Table 3 shows the comparison of the previous procedures (entries [1][2][3][4][5][6][7][8][9] used for the synthesis of 3,4-Dihydro-3,3-dimethyl-13-(4-methylphenyl)-2H-indazolo[2,1-b]phthalazine-1,6,11(13H)-trionewith our used method (entry 10). In general, each procedure includes some advantages that by focusing on the pervious routes we could declare this method could accelerate the process of reaction and be useful and e cient as well. To investigate the recyclability of the Mn supported nano-catalyst for environmental and commercial applications, it was tested with the model reaction, and found that it is relatively capable even after ve runs and its activity was almost similar to the fresh one (90, 90, 88, 87, and 84%) respectively (Fig. 8). The stability of the Mn supported nano-catalyst even after the 5th run was inspected by FT-IR and SEM techniques ( Fig. 9-10).

Antibacterial properties
The antibacterial properties of the Mn supported nano-catalyst were studied against a number of grampositive and gram-negative bacterial strains, and DMSO was used as a blank ( Table 4). The catalyst inhibited the growth of bacterial strains, producing a zone of inhibition of diameter 5-30 mm. The Mn supported nano-catalyst was even more effective against Serratia marcescens in gram-negative bacteria than the standard tetracycline antibiotic. Since the comparison of the size of inhibition zones is generally not reliable, the MIC value of the compound was also determined. The results indicated that the MIC value of the Mn supported nano-catalyst against the tested organisms was about 8 mg/mL. The MIC value of standard tetracycline antibiotic is about 8 mg/mL.

Conclusion
In conclusion, we were successful to synthesize a novel Mn nano-catalyst supported on magnetic iron oxide functionalized with mercaptobenzoxazole as an appropriate surface for synthesis of 2H-Indazolo[1,2-b]phthalazine-triones from the one pot three-component condensation reaction of aldehydes, dimedone and phthalhydrazide in re uxed ethanol in ne yields and short reaction times. In fact iron oxide nanoparticles through the noticeable properties such as small size, high magne-tism and low toxicity would be a proper methods for chemistry synthesis. Also we inspected the antibacterial properties of Mn nano catalyst against a number of gram-positive and gram-negative bacterial strains and determined the nano catalyst would be more effective against Serratia marcescens in gram-negative bacteria than the standard tetracycline antibiotic. Environmental issues revealed that the Mn nano catalyst would be used for ve runs without signi cant loss of the catalytic activity.   The EDX analysis of the Mn supported nano-catalyst Figure 6