Preparation, Antibacterial Activity, and Catalytic Application of Magnetic Graphene Oxide‐Fucoidan in the Synthesis of 1,4‐Dihydropyridines and Polyhydroquinolines

1 Introduction

Natural biopolymers have aroused renewed interest as an efficient tool in the development of biodegradable materials.1-3 Their particularly attractive and desirable properties including nontoxic nature, biocompatibility, and biodegradability along with plentiful sources amongst natural materials make them prospective materials for many uses.4 Recently, carbohydrate-based marine polymers have been widely used in the construction of nanocomposites, due to their high number of surface functionalities and biocompatibility, making them useful as heterogeneous catalysts.5-7 Fucoidan (Fu), one of the most substantial cost-effective marine polysaccharides originating from brown algae, is a fucose-rich hygroscopic sulfated polysaccharide containing negative charges, due to its sulfate functional groups, which allow it to form complexes with other molecules of opposite charge.8, 9 Moreover, Fu, built of a backbone of α(1→3)-l-fucopyranose residues having various substituents, has attracted great attention as capping material to design new nanocomposites. Besides, outstanding pharmaceutical applications of fucoidan including antibacterial, anticoagulant, immune, and anti-thrombotic effects, have been widely studied and established.3, 10, 11

Nanocomposite technology is considered an effective strategy in materials research and development of polymer nanocomposites by using low-loaded fillers, such as carbon nanomaterials (carbon nanotubes and graphene sheets), clay, hydroxyapatite, etc. Among these, graphene oxide (GO) with a remarkable amount of oxygenated groups such as hydroxyl, epoxide, and carboxyl groups provides effective means to alter the interactions between polymer chains and GO sheets and the reinforcement of nanocomposites.12-15 However, for the separation of GO, a high-speed centrifuge is needed, which would discourage its application. Nevertheless, this disadvantage can be eliminated using magnetic GO which can be easily separated from the solution by a magnet.

During recent years, metal nanoparticles especially magnetic nanoparticles (MNPs) as a solid support material for the development of magnetically retrievable catalytic systems have attracted attention due to the easy separation, high surface area, accessibility, high level of chemical and thermal stability, and recoverability which have been considerably developed.16-19 Moreover, MNPs have gained increasing attention due to their promising applications of including biomolecular sensing, biomedical applications, pigments, and heterogeneous catalysis. Among them, barium hexaferrite (BaFe12O19), M-type permanent magnet, showed promising applications in heterogeneous catalysis and microwave absorbing materials.20, 21

Considering the above-mentioned facts, in this work, we have designed and developed a new magnetic heterogeneous catalyst using a natural sulfated polysaccharide, Fucoidan, for the functionalization of GO magnetized by barium ferrite named BaFe12O19@GO@Fu providing a unique combination of excellent properties.

Interesting features of this catalyst including the synergistic effect of Fucoidan with graphene oxide which improved its catalytic capacity in different conditions due to the presence of functional groups on the surface of both materials.22 In the other words, not only the negative charge of Fucoidan can catalyze the reactions demanding a basic catalyst.23 but also, the presence of graphene oxide because of carboxylic acid and proton donor groups on its surface enables to catalyze the reactions in which need to acidic catalysis, and besides the bi-functional perspective of this part of the catalyst, thanks to the barium ferrite with noticeable magnetic properties make catalyst separation convenient at the end of the reaction. Finally, the high efficiency of this catalyst in the synthesis of 1,4-dihydropyridines and polyhydroquinolines, is worth noting.

Nowadays, numerous structural analogs of these compounds are used in pharmacy and medicine.24 For instance, amlodipine, nicardipine, and nifedipine belong to the dihydropyridine family (Figure 1a).25 The classical approach for the synthesis of these compounds is one-pot condensation reactions of various aldehydes with β-ketoesters, dimedone, and ammonium acetate under drastic conditions. In recent decades, some improved procedures using an extensive range of catalysts including magnetic chitosan-terephthaloyl-creatine,26 BiFeO3,27 and γ-Fe2O3/Cu@cellulose,28 Keggin-type heteropolyacid H5BW12O40,29 and amine-functionalized graphene oxide nanosheets (AFGONs)13 have been reported in the literature. While many of them have their merits, some of them have a variety of drawbacks, including harsh reaction conditions, expensive catalysts, and a tedious preparation and work-up process. Thus, the design and expansion of an alternate approach for this reaction is a priority. Given the importance of 1,4-dihydropyridine and polyhydroquinoline, and in the framework of our previous research on the use of environment-friendly natural polysaccharides in the synthesis of potential biologically active compounds.3, 6, 7, 30-32 We report herein the catalytic activity investigation of BaFe12O19@GO@Fu in the synthesis of biologically active and pharmaceutically important 1,4-dihydropyridine and polyhydroquinoline under reflux in ethanol (Figure 1b).

a) Examples of 1,4-dihydropyridine pharmaceutical derivatives, b) BaFe12O19@GO@Fu catalyzed green synthesis of 1,4-dihydropyridines 5 a–e and polyhydroquinoline 6 f–n

In addition to the catalytic investigation, the antibacterial activity of BaFe12O19@GO@Fu was evaluated on two bacterial strains, gram-negative Escherichia coli (E. coli) and gram-positive Staphylococcus aureus (S. aureus).

2 Results and Discussion

The FTIR spectra of BaFe12O19, GO, Fu, and BaFe12O19@GO@Fu nanocomposite are shown in Figure 2. In the FTIR spectrum of BaFe12O19@GO@Fu, the peaks at 403 and 576 cm−1 are related to the metal-oxide stretching vibration from the BaFe12O19 structure. Strong broadband at 3500 cm−1 is associated with the stretching vibration due to the O−H of Fucoidan. A bond at 1670 cm−1 can be attributed to representative polysaccharide chains. The absorption band at 993 cm−1 indicated hemiacetal vibration at alcohol and ether functional groups in the Fucoidan structure. The peak at 1249–1431 cm−1 is related to the stretching vibration of S=O from the SO3H group. Also, in the IR spectrum of GO, the absorption band located at 3000–3600 cm−1 corresponds to hydrogen-bonded O−H stretch, and the peak at 1731 cm−1 is related to the C=O stretching. The bending vibration of OH appears at 1614 cm−1 and the peak at 1091 cm−1 corresponds to the vibrational mode of the C−O group. The shift of OH stretching vibrations to lower wavenumber indicates the increase in intermolecular hydrogen bonding between GO and Fu. It can be seen that all the characteristic peaks of the constituents, namely GO, Fu, and BaFe12O19, are present in the IR spectrum of the nanocomposite.

FTIR spectra of BaFe12O19, GO, Fu, BaFe12O19@GO@Fu

Magnetic measurements were carried out using a vibrating sample magnetometer (VSM) at room temperature by using a magnetic field ranging from −10000 to 10000 Oe (Figure 3a). The remnant magnetization value of BaFe12O19 nanoparticles was about 57.04 emu g−1 implying the ferrimagnetic behavior of nanoparticles. According to the magnetization curve of the BaFe12O19@GO@Fu nanocomposite, the value of the saturation magnetization was 18.28 emu g−1. The smaller value of the saturation magnetization of BaFe12O19@GO@Fu compared to BaFe12O19 is due to the nonmagnetic layer-by-layer surface coverage.

a) VSM magnetization curve of BaFe12O19, BaFe12O19@Go@Fu, and b) TGA curves of Fu and BaFe12O19@GO@Fu nanocomposite

The thermal behavior of the prepared nanocomposite was probed by TGA analyses at the range of temperature between 50 and 800 °C with the rate of 10 °C min−1. Figure 3b shows the TGA curves of Fucoidan and BaFe12O19@GO@Fu nanocomposite. As illustrated in the TGA curve of Fu, the majority of weight loss within a range of 200–400 °C indicated the thermal degradation of the polysaccharide. From the TGA curve of BaFe12O19@GO@Fu, it is clear that a tiny fraction of weight reduction occurred upon heating at about 100–200 °C, which relates to the elimination of volatile elements such as water from the nanocomposite. After that, the mass loss in the range of 230–335 °C to approximately 8 % and 430–500 °C just below 12 % of nanocomposite can be associated with the decomposition of GO and Fu. It should be emphasized that the BaFe12O19@GO@Fu magnetic nanocomposite shows higher thermal stability.

Initially, the size, structure, and morphology of the BaFe12O19 NPs and of the BaFe12O19@GO@Fu nanocomposite were investigated by SEM analysis (Figures 4a–d), which is testimony to the fact that BaFe12O19 nanoparticles have a completely uniform M-type structure. Furthermore, the structure and size of the nanoparticles at 10, 1 μm, and 500 nm were monitored by SEM analysis which confirms the preservation of morphology and particle size (Figures 4a–d).

SEM images of a) BaFe12O19, b–d) BaFe12O19@GO@Fu (10 μm, 1 μm, 500 nm), and e) The EDX analysis of the BaFe12O19@GO@Fu magnetic nanocomposite.

In fact, the prepared nanoparticles were uniformly loaded on the composite surface and the averaged size for these nanoparticles was approximately 53–76 nm. Also, the presence of S, O, C, Ba, and Fe elements in the studied composite was confirmed by using EDX analysis, and the presence of these elements could be attributed to barium hexaferrite, graphene oxide, and fucoidan (Figure 4e).

X-ray diffraction patterns of the bare BaFe12O19, GO@Fu, and BaFe12O19@Go@Fu are shown in Figure 5. The diffraction peaks observed at 2θ values of 29.92°, 31.68°, 33.54°, 34.84°, 36.54°, 54.61°, 56.18°, and 62.84° correspond to the crystal planes (110), (107), (114), (203), (217), (2011), (220) and confirmed that BaFe12O19 nanoparticles were synthesized in hexagonal crystal system based on the standard XRD pattern (JCPDS, card number 01–072-0738). According to the Scherrer equation, the size of the nanoparticles amounted to 85 nm.

X-ray diffraction patterns of a) the bare BaFe12O19 b) GO@Fu and c) BaFe12O19@GO@Fu.

The XRD pattern of GO@Fu indicates a sharp peak at about 2θ≈12° which is related to graphene oxide and another peak at about 2θ≈27° which can refer to the amorphous nature of Fu (Figure 5b). The XRD pattern of BaFe12O19@GO@Fu nanocomposite indicates that the crystal structure of barium hexaferrite has been conserved after modification, and at the same time, the existence of a small bump in 2θ about 12° and 27° might confirm the presence of GO and Fu in the nanocomposite (Figure 5c).

The shape and size of the nanoparticles BaFe12O19@Go@Fu were examined by Transmission Electron Microscopy (TEM) analysis. TEM images with different magnifications (300 and 200) reveal the layered nature of the composites and a random distribution of the rod-shaped BaFe12O19 nanoparticles (black parts) in the GO sheets (white parts) which approves the interactions between nanoparticles and GO surface (Figure 6).

TEM images of BaFe12O19@GO@Fu nanocomposites at different magnification.

The BaFe12O19@GO@Fu nanocomposite was characterized by Raman spectroscopy analysis as shown in Figure 7. The D band of GO is found at 1351 cm−1 which is associated with the disorder stemming from oxygen moieties and the G band is found at 1593 cm−1 due to C−C stretching. The observed intensity ratio (ID/IG) was 0.87 for multilayer GO. The Raman spectrum also shows some characteristic peaks of the M-type barium ferrite phase. The peaks at 758 and 684 cm−1 can be assigned to A1g vibrations of Fe−O bonds at the tetrahedral 4f1 and bipyramidal 2b sites, respectively.34 The peaks at 299 cm−1 are due to E1g vibrations, while the peak at 342 cm−1 is due to E2g vibration. The Raman band at 840 cm−1 is attributed to the COS bending vibration of the sulfate group in fucoidan.

Raman spectrum of BaFe12O19@GO@Fu.

2.1 Antibacterial activity of BaFe12O19@GO@Fu

The antibacterial efficacy of BaFe12O19@GO@Fu was investigated by the Agar well diffusion method (Figure 8). A suspension of bacteria (E. coli or S. aureus) was diffused on the agar plates of Mueller-Hinton (MH) supplemented with Tween 80 surfactants (final concentration of 0.05 % v/v) using a final density of 1.5×108 colony-forming units (CFU)/ml of tested bacterial strains suspended in MH. Subsequently, on the surface of the previously inoculated agar plate, 50 mg of sample was placed which was firstly kept at 4 °C for 2 h, then incubated at 37 °C for 24 h. The clear zones of inhibition (13 mm) around the antimicrobial sample revealed that BaFe12O19@GO@Fu has antibacterial activity against gram-positive bacteria S. aureus.

Antibacterial activity of BaFe12O19@GO@Fu against a) E. coli and n) S. aureus.

In connection with the mechanism of this inhibitn, studies to date have not yet determined the exact mechanism for antibacterial activity, but the possible mechanism could be based on the binding of sulfate groups in fucoidan to the bacterial cell wall resulting in bacterial cell membrane destruction, and leakage of intracellular material to the outside, eventually causing the death of the bacteria.

2.2 Catalytic Activity of BaFe12O19@GO@Fu Magnetic Nanocomposite

The catalytic efficacy of BaFe12O19@GO@Fu was investigated in the one-pot reaction between 4-chlorobenzaldehyde (1 mmol), ethyl acetoacetate (1 mmol), dimedone (1 mmol), and ammonium acetate (2 mmol) as a model reaction for the synthesis of polyhydroquinoline derivatives. In the case of 1,4-dihydropyridines, ethyl acetoacetate (2 mmol) has been used instead of dimedone. Both of the model reactions were performed in the presence of different solvents such as ethanol, water, toluene, dichloromethane, acetonitrile, and also under solvent-free conditions. The results show a substantial increase in the yield of the reaction when ethanol was used as a solvent. To find the optimum catalyst loading, the model reactions were studied, regarding various amount of catalyst (Table 1). Although the steady increase in the amount of catalyst from 5 to 15 mg (entries 7 to 10) gives noticeable yield, a further quantity of catalyst does not give significant change in the yield of the reaction. Model reactions were carried out in the absence of catalyst (entry 1) and with Fu, BaFe12O19, GO@Fu, and BaFe12O19@GO@Fu (entries 2 to 5). These results endorsed that BaFe12O19@GO@Fu was more appropriate for these reactions. Overall, the most influential conditions for the desired products were found to be refluxing in ethanol in the presence of 15 mg magnetic nanocomposite.

Table 1. Optimization of the catalyst, temperature, and the solvent for the synthesis of 1,4-dihydropyridine and polyhydroquinolines.

Product 5d[b]

Product 6 m[a]

Solvent

Temperature

Catalyst (amount in [mg])

Entry

Yield (%)

Time (min)

Yield (%)

Time (min)

trace

300

trace

300

Solvent free

r.t.

–

1

78

20

80

20

Ethanol

Reflux

GO (15)

2

61

20

65

20

Ethanol

Reflux

BaFe12O19 (15)

3

80

20

85

20

Ethanol

Reflux

Fu (15)

4

85

15

89

10

Solvent free

r.t.

BaFe12O19@GO@Fu (15)

5

90

15

91

10

Ethanol

r.t.

BaFe12O19@GO@Fu (15)

6

90

15

90

12

Ethanol

Reflux

BaFe12O19@GO@Fu (5)

7

91

15

93

12

Ethanol

Reflux

BaFe12O19@GO@Fu (10)

8

95

15

95

12

Ethanol

Reflux

BaFe12O19@GO@Fu (12)

9

96

15

97

12

Ethanol

Reflux

BaFe12O19@GO@Fu (15)

10

90

15

91

12

Ethanol

Reflux

BaFe12O19@GO@Fu (20)

11

78

20

80

20

H2O

Reflux

BaFe12O19@GO@Fu (15)

12

90

15

91

15

Ethanol/H2O

Reflux

BaFe12O19@GO@Fu (15)

13

60

30

60

30

CH3CN

Reflux

BaFe12O19@GO@Fu (15)

14

55

35

60

35

Toluene

Reflux

BaFe12O19@GO@Fu (15)

15

50

45

55

45

CH2Cl2

Reflux

BaFe12O19@GO@Fu (15)

16

[a] Reaction conditions: 4-chlorobenzaldehyde (1 mmol), ethyl acetoacetate (1 mmol), dimedone (1 mmol), ammonium acetate (1 mmol), and 3 ml solvent; [b] reaction conditions: 4-chlorobenzaldehyde (1 mmol), ethyl acetoacetate (2 mmol), ammonium acetate (2 mmol), and 3 ml solvent.

For overall evaluation of the synthesis of 1,4-dihydropyridines and polyhydroquinolines after the aforementioned optimization of conditions, a range of various aromatic aldehydes were chosen. As can be seen in Table 2, substrates with both electron-donating and electron-withdrawing substituents were investigated in these reactions. The presence of electron-withdrawing substituents tended to increase the reaction rate, and the electron-donating group slowed down the process. The results clearly show that the reaction of diverse aromatic aldehydes, ammonium acetate, ethyl acetoacetate, and dimedone under reflux with the presence of the nanocomposite provided the corresponding products in high yields (90–96 %) at appropriate reaction times. As most of the products crystallized directly from the reaction mixture, all of the products were characterized by their melting points, and some of the products were additionally characterized by NMR spectral data (s. Supporting Information).

Table 2. Synthesis of 1,4-dihydropyridine and polyhydroquinolines catalyzed by BaFe12O19@GO@Fu nanocomposite.

Entry

Product

Aldehyde

Time [min]

Yield [%]

M.p. [°C] Found/Reported

1