Journal of Chemistry

Volume 2014 (2014), Article ID 457430, 8 pages

http://dx.doi.org/10.1155/2014/457430

Research Article

Synthesis, Single Crystal X-Ray Structure, and Antimicrobial Activity of 6-(1,3-Benzodioxol-5-ylmethyl)-5-ethyl-2-{[2-(morpholin-4-yl)ethyl]sulfanyl}pyrimidin-4(3H)-one

Mohamed I. Attia,1,2 Nasser R. El-Brollosy,1,3 Amany L. Kansoh,⁴ Hazem A. Ghabbour,¹ Reem I. Al-Wabli,¹ and Hoong-Kun Fun¹

¹Department of Pharmaceutical Chemistry, College of Pharmacy, King Saud University, P.O. Box 2457, Riyadh 11451, Saudi Arabia

²Pharmaceutical and Drug Industries Research Division, Medicinal and Pharmaceutical Chemistry Department, National Research Centre, Dokki, Giza 12622, Egypt

³Department of Chemistry, Faculty of Science, Tanta University, Tanta 31527, Egypt

⁴Genetic Engineering and Biotechnology Division, Microbial Chemistry Department, National Research Centre, Dokki, Giza 12622, Egypt

Received 8 April 2014; Accepted 8 May 2014; Published 26 May 2014

Academic Editor: Hakan Arslan

Copyright © 2014 Mohamed I. Attia et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract.

Synthesis, X-ray structure and antimicrobial activity of 6-(1,3-benzodioxol-5-ylmethyl)-5-ethyl-2-{[2-(morpholin-4-yl)ethyl]sulfanyl}pyrimidin-4(3H)-one (8) are reported. Compound 8 exhibited activity towards S. aureus with MIC value of 0.0619 μmol/mL while it showed activity towards B. subtilis, B. cereus, C. albicans, and A. niger with MIC = 0.1859 μmol/mL. Single crystal X-ray structure of the title compound 8 confirmed its S-alkylation. The title compound crystallizes in the triclinic, -1, (5) Å, (5) Å, (9) Å, (2)°, (2)°, (2)°, (2) Å³, , , ,  K. The crystal structure is stabilized by weak intermolecular C–HO and N–HO hydrogen interactions.

1. Introduction

Pyrimidines are attractive scaffolds for drug design and development since their discovery as components of DNA and RNA. Besides, pyrimidines are incorporated in a plethora of bioactive molecules with various biological activities. The chemotherapeutic effect of pyrimidines could be attributed to their ability to inhibit vital enzymes responsible for the nucleic acid biosynthesis. Several pyrimidines showed antiviral [1–3], antimicrobial [4–8], anticancer [9–11], anti-inflammatory [12, 13], antileishmanial [14, 15], and cardiovascular activities [16, 17].

On the other hand, a sizable number of bioactive molecules bearing 1,3-benzodioxole moiety exhibited multiple biological activities like antioxidant [18, 19], anticancer [20, 21], anti-inflammatory [22, 23], anticonvulsant [24], antihypertensive [25], antidepressant [26], immunomodulators [27], and antiprotozoal [28, 29].

In continuation of our ongoing research on the synthesis of potential antimicrobial pyrimidinones [30], we report herein the synthesis, single crystal X-ray structure, and antimicrobial activity of new pyrimidinone derivative incorporating 1,3-benzodioxole moiety, namely, 6-(1,3-benzodioxol-5-ylmethyl)-5-ethyl-2-{[2-(morpholin-4-yl)ethyl]sulfanyl}pyrimidin-4(3H)-one.

2. Experimental

2.1. General

Melting points were determined on a Gallenkamp melting point apparatus and are uncorrected. Silica gel TLC (thin layer chromatography) plates from Merck (silica gel precoated aluminium plates with fluorescent indicator at 254 nm) were used for thin layer chromatography. Visualization was performed by illumination with UV light source (254 nm). Compound 4 was prepared according to the reported literature procedures [31–33]. The tested microorganisms (Gram-positive bacteria, Staphylococcus aureus ATCC 29213, Bacillus subtilis NRRL 4219, and Bacillus cereus; pathogenic fungi, Candida albicans ATCC 10231 and Aspergillus niger NRRL 599) were obtained from MIRCIN Cairo, Faculty of Agriculture, Ain Shams University, Cairo, Egypt. The reference drugs ampicillin trihydrate (CAS 7177-48-2) and clotrimazole (CAS 23593-75-1) were obtained from Sigma-Aldrich Chemie GmbH (Taufkirchen, Germany). The X-ray diffraction measurements of compound 8 were performed using Bruker Venture D8 diffractometer.

2.2. Synthesis of 6-(1,3-Benzodioxol-5-ylmethyl)-5-ethyl-2-thioxo-2,3-dihydropyrimidin-4(1H)-one (7)

A suspension of activated zinc dust [30] (7 g) in dry THF (30 mL) was heated to reflux. Few drops of ethyl 2-bromobutanoate (5) were added and the reaction mixture was refluxed for 10 min. The piperonylcarbonitrile (4) (2.42 g, 0.015 mol) was added in one portion while the rest of ester 5 (5.85 g, 0.03 mol) was added dropwise. After complete addition, the reaction mixture was refluxed for 30 min, then diluted with THF (100 mL), and quenched by addition of sat. aq. K₂CO₃ solution (30 mL). The reaction mixture was stirred for 1 h at room temperature and then the THF layer was removed and the residue was washed with THF (3 × 20 mL). The combined THF fractions were stirred with 10% aq. HCl (20 mL) for 30 min. The solution was concentrated under reduced pressure and diluted with CH₂Cl₂ (100 mL). The organic phase was washed with sat. aq. NaHCO₃ solution (2 × 50 mL), dried (Na₂SO₄), and evaporated under reduced pressure to give the respective β-ketoester 6 as a pale yellow viscous oil. The crude β-ketoester 6 (2.78 g, 0.01 mol) was added to a boiling solution of sodium ethoxide (prepared from 4.9 g of Na in 100 mL of absolute EtOH) containing thiourea (11.42 g, 0.15 mol). The reaction mixture was heated to reflux for 24 h. The solvent was evaporated to dryness under reduced pressure and the residue was dissolved in H₂O (100 mL). The product was precipitated by addition of conc. HCl till pH = 4. The precipitate was filtered off, washed with H₂O, and dried to give the corresponding thiouracil derivative 7. The crude 7 was purified by recrystallization (ethanol) to afford pure 7 in 46% yield as a white solid mp 183–185°C.

2.3. Synthesis of 6-(1,3-Benzodioxol-5-ylmethyl)-5-ethyl-2-{[2-(morpholin-4-yl)ethyl]sulfanyl}pyrimidin-4(3H)-one (8)

Anhydrous potassium carbonate (0.304 g, 2.2 mmol) was added to a solution of thiouracil 7 (0.29 g, 1 mmol) in dry DMF (5 mL) followed by addition of N-(2-chloroethyl) morpholine hydrochloride (0.21 g, 1.1 mmol). The reaction mixture was stirred at room temperature for 24 h; then the solvent was evaporated under reduced pressure. The residue was chromatographed on silica gel column with CHCl₃ to afford the corresponding 2-[2-(morpholin-4-yl)ethyl]thiouracil derivative 8 in 58% yield as a white solid mp 138–140°C.

2.4. Crystal Structure Determination

Slow evaporation of the pure compound 8 in dimethyl sulfoxide yielded its colourless single crystals. A colourless block-shaped single crystal of suitable size, 0.65 × 0.32 × 0.19 mm, was selected for X-ray diffraction analysis. Data were collected on a Bruker Venture D8 CMOS area diffractometer equipped with graphite monochromatic radiation ( Å) at 100 K. Cell refinement and data reduction were done by Bruker SAINT [34]; program used to solve structure and refine structure is SHELXTL [35]. The final refinement was performed by full-matrix least-squares techniques with anisotropic thermal data for non hydrogen atoms on . All the hydrogen atoms were placed in calculated positions and constrained to ride on their parent atoms. Multiscan absorption correction was applied by use of SADABS software [34]. The crystallographic data and refinement information are summarized in Table 1.

Table 1: Crystallographic data and refinement information.

Empirical formula
C₂₀H₂₅N₃O₄S

Formula weight
403.49

Temperature (K)
100

Crystal system
Triclinic

Space group
P-1

Mo radiation, λ
0.71073 Å

(Å)
11.1220 (5)

(Å)
12.2241 (5)

(Å)
21.5246 (9)

(°)
88.958 (2)

(°)
79.836 (2)

(°)
79.384 (2)

(Å³)
2830.9 (2)

6

F(000)
1284

θ range (°)
2.3–28.0

(mm⁻¹)
0.21

Density (calc.) (g/cm³)
1.420

Crystal shape and color
Block, colorless

Crystal size (mm³)
0.65 × 0.32 × 0.19

h/k/l
−14,14/−16,16/−28,28

Measured reflections
86637

Independent reflections
14111

Reflections with
9629

Goodness-of-fit on
1.01

[ > 2σ()]
0.046

wR
0.110

(e Å⁻³)
0.37

(e Å⁻³)
−0.33

2.5. Antimicrobial Activity by the Agar Disc-Diffusion Method

Sterile nutrient, malt extract, and Czapek’s dox agar media were inoculated, separately, with 100 μL cell suspension of the chosen bacteria, Candida albicans, and Aspergillus niger, respectively, and poured into Petri dishes (20 cm diameter). The test compound 8 (200 μg/10 mm diameter disc) was placed onto the surface of the agar Petri dishes. The antimicrobial potential was expressed as the diameter of the growth inhibition zone in mm [36].

2.6. Minimum Inhibitory Concentrations (MICs)

The minimum inhibitory concentration (MIC) of the test compound 8 was evaluated using serial dilutions technique [37]. Different concentrations ranging from 50 to 200.0 μg for the test compound 8 were dissolved in dimethyl sulfoxide (1 mL) and were placed on filter paper disc (1 cm diameter). The discs were deposited on the surface of inoculated agar plates and kept at low temperature before incubation which favours diffusion over microbial growth to detect the inhibition zone clearly. The plates were incubated at 30°C for 24 h for bacteria and Candida albicans and for 48 h for Aspergillus niger.

3. Results and Discussion

3.1. Chemistry

The commercially available piperonal (1) was elaborated to piperonylcarbonitrile (4) as portrayed in Scheme 1. Thus, piperonal (1) was reduced with sodium borohydride in methanol at room temperature to give the piperonyl alcohol (2) [31] in almost quantitative yield. The alcohol 2 was allowed to react with thionyl chloride to give the chloro derivative 3 [32]. Subsequently, compound 3 was subjected to nucleophilic substitution using sodium cyanide and a catalytic amount of potassium iodide in dimethylformamide to yield the respective piperonylcarbonitrile (4) [33] in about 75% overall yield.

Scheme 1: Synthesis of the intermediate piperonylcarbonitrile (4). Reagents and conditions: (i) NaBH₄, methanol, RT, 18 h; (ii) SOCl₂, RT, 18 h; (iii) NaCN, KI, DMF, RT, 18 h.

Scheme 2 illustrates the synthetic pathway to achieve the target compound 8 and its intermediates. Compound 4 was allowed to react with ethyl 2-bromobutanoate (5) in dry tetrahydrofuran in the presence of zinc dust under Reformatsky conditions [38] to furnish ethyl 4-(1,3-benzodioxol-5-yl)-2-ethyl-3-oxobutanoate (6) in good yield. Reaction of thiourea with the crude β-ketoester 6 in absolute ethanol in the presence of sodium ethoxide gave the pivotal 6-(1,3-benzodioxol-5-ylmethyl)-5-ethyl-2-thioxo-2,3-dihydropyrimidin-4(1H)-one (7) in 46% yield. Thiouracil 7 was reacted with N-(2-chloroethyl) morpholine hydrochloride in the presence of anhydrous potassium carbonate in dimethylformamide at room temperature to afford the title compound 6-(1,3-benzodioxol-5-ylmethyl)-5-ethyl-2-{[2-(morpholin-4-yl)ethyl]sulfanyl} pyrimidin-4(3H)-one (8) in 58% yield. Single crystal X-ray crystallography confirmed the 3D structure of compound 8.

Scheme 2: Synthesis of the target compound 8. Reagents and conditions: (i) (1) Zn/THF, (2) K₂CO₃, and (3) HCl; (ii) (1) NH₂CSNH₂, NaOEt, ethanol, reflux, 24 h and (2) HCl; (iii) N-(2-chloroethyl) morpholine hydrochloride, K₂CO₃, DMF, RT, 18 h.

3.2. Crystal Structure of Compound 8

Single crystal X-ray crystallography is a doubtlessly decisive analytical tool which can confirm the 3D structure of the target compound 8, particularly its S-alkylation. Fortunately, we have succeeded to get single crystals of compound 8 which were suitable for X-ray crystallography (Figure 1).

Figure 1: ORTEP diagram of the title compound 8 drawn at 50% ellipsoids for non hydrogen atoms.

The asymmetric unit contains three molecules of the title compound 8. The molecular packing in the crystal structure is stabilized by the weak intermolecular interactions (Figure 2, Table 2), of which O3B, O2A, and O4B work as acceptor while N2B, C3A, and C19B work as donors. The distance H1NBO3B is 1.87 (2) Å, H3ABO2A is 2.5300, and H19DO4B is 2.5600 Å, and the angles between N2B–H1NBO3B, C3A–H3ABO2A, and C19B–H19DO4B are 173 (2), 146.00 (2), and 140.00° (2), respectively. There are four intramolecular H-bonds in the molecules: in the first molecule, there is one H-bond between C16A–H16BN1A; in the second molecule, there is one H-bond between C15B–H15CN1B; and in the third molecule, there are two H-bonds between C16C–H16EN1C and C17C–H17FO1C. The selected bond lengths, bond angles, and bond torsion angles are listed in Table 3.

Table 2: Hydrogen-bond geometry (Å, °).

D–HA
D–H
HA
DA
D–HA

N2C–H1NCO3A
0.92 (2)
1.85 (2)
2.7721 (18)
178.0 (12)

N2B–H1NBO3Bi
0.92 (2)
1.87 (2)
2.7904 (19)
173 (2)

N2A–H1NAO3C
0.97 (2)
1.76 (2)
2.7330 (18)
179 (3)

C3A–H3ABO2Aii
0.9700
2.5300
3.371 (2)
146.00

C15B–H15CN1B
0.9700
2.4800
2.882 (2)
105.00

C16A–H16BN1A
0.9700
2.6100
3.198 (2)
119.00

C16C–H16EN1C
0.9700
2.5800
3.170 (2)
120.00

C17C–H17FO1C
0.9700
2.5900
3.441 (2)
146.00

C19A–H19BO2B
0.9700
2.5000
3.462 (2)
172.00

C19B–H19DO4Biii
0.9700
2.5600
3.366 (3)
140.00

Symmetry codes: (i) ; (ii) ; (iii) .

Table 3: Selected geometric parameters (Å, °) for one molecule.

S1A–C14A
1.7546 (18)
O3C–C13C
1.244 (2)

S1A–C15A
1.8062 (18)
O4C–C18C
1.431 (2)

O1A–C2A
1.377 (2)
N2A–C13A
1.380 (2)

O1A–C3A
1.421 (3)
N3A–C17A
1.463 (2)

O2A–C4A
1.383 (2)
N3A–C20A
1.469 (2)

O2A–C3A
1.432 (3)
N3A–C16A
1.466 (2)

O3A–C13A
1.241 (2)
N1B–C9B
1.380 (2)

O4A–C18A
1.425 (2)
N1B–C14B
1.295 (2)

O4A–C19A
1.425 (2)
N2B–C14B
1.365 (2)

C14A–S1A–C15A
101.81 (8)
N3A–C17A–C18A
110.66 (17)

C2A–O1A–C3A
105.29 (15)
N3A–C20A–C19A
109.97 (14)

C3A–O2A–C4A
104.85 (15)
O1B–C2B–C1B
128.33 (17)

C18A–O4A–C19A
109.78 (13)
O1B–C2B–C4B
109.39 (15)

C9A–N1A–C14A
116.61 (14)
O3B–C13B–N2B
119.86 (15)

C13A–N2A–C14A
122.13 (14)
N2B–C13B–C10B
115.59 (15)

C16A–N3A–C17A
108.97 (14)
S1B–C14B–N1B
121.81 (13)

C17A–N3A–C20A
108.05 (13)
N1B–C14B–N2B
124.00 (16)

C16A–N3A–C20A
110.73 (13)
S1B–C14B–N2B
114.19 (12)

O1A–C2A–C4A
110.00 (15)
O2C–C4C–C2C
109.89 (15)

O1A–C2A–C1A
127.51 (18)
N1C–C9C–C8C
112.44 (14)

O1A–C3A–O2A
108.23 (15)
N1C–C9C–C10C
123.70 (15)

O2A–C4A–C2A
109.69 (16)
O3C–C13C–C10C
124.24 (16)

O2A–C4A–C5A
128.57 (18)
O3C–C13C–N2C
119.71 (15)

N1A–C9A–C10A
123.90 (15)
N2C–C13C–C10C
116.05 (15)

N1A–C9A–C8A
112.67 (14)
S1C–C14C–N2C
113.52 (12)

O3A–C13A–N2A
119.66 (15)
S1C–C14C–N1C
122.83 (13)

N2A–C13A–C10A
115.36 (15)
N1C–C14C–N2C
123.65 (16)

O3A–C13A–C10A
124.97 (16)
S1C–C15C–C16C
112.16 (13)

S1A–C14A–N1A
122.83 (13)
N3C–C16C–C15C
113.16 (15)

S1A–C14A–N2A
113.20 (12)
N3C–C17C–C18C
111.34 (16)

N1A–C14A–N2A
123.97 (16)
O4C–C18C–C17C
111.12 (14)

S1A–C15A–C16A
111.98 (13)
O4C–C19C–C20C
111.83 (16)

N3A–C16A–C15A
112.40 (15)
N3C–C20C–C19C
110.99 (14)

Figure 2: Crystal packing showing intermolecular C–HO hydrogen bonds as dashed lines.

3.3. In Vitro Antimicrobial Activity

The in vitro antimicrobial activity of compound 8 was evaluated against Gram-positive bacteria (S. aureus, B. subtilis, and B. cereus) and pathogenic fungi (C. albicans and Aspergillus niger). The obtained data, expressed as diameter of the inhibition zone (DIZ) and minimum inhibitory concentration (MIC) for the test compound 8 as well as for the reference antibacterial standard, ampicillin, and the reference antifungal standard, clotrimazole, are shown in Tables 4 and 5.

Table 4: Diameter of growth inhibition zone (mm) of the target compound 8, ampicillin, and clotrimazole towards the test microorganisms.

Compound number
Diameter of growth inhibition zone ± SD (mm)*

Gram-positive bacteria
Pathogenic fungi

Staphylococcus aureus
Bacillus subtilis
Bacillus cereus
Candida albicans
Aspergillus niger

8
20 ± 0.50
18 ± 1.00
19 ± 1.50
17 ± 1.50
20 ± 1.50

Ampicillin
32 ± 1.00
28 ± 1.00
27 ± 0.50

Clotrimazole

35 ± 1.00
32 ± 0.00

The arithmetic mean of the inhibition zone diameters in mean ± standard deviation in mm.

Table 5: The minimum inhibitory concentration (MIC, mol/mL) values for the target compound 8, ampicillin, and clotrimazole towards the test microorganisms.

Compound number
The minimum inhibitory concentrations (MICs, mol/mL)*

Gram-positive bacteria
Pathogenic fungi

Staphylococcus aureus
Bacillus subtilis
Bacillus cereus
Candida albicans
Aspergillus niger

8
0.0619
0.1859
0.1859
0.1859
0.1859

Ampicillin
0.0258
0.0258
0.0258

Clotrimazole

0.0261
0.0348

The lowest concentration of the test compound that inhibits the growth of microorganism (mol/mL).

The preliminary antimicrobial potential of the test compound 8 was evaluated using DIZ assay and the results are given in Table 4. Compound 8 exhibited good antimicrobial activity with DIZ values in the range of 17–20 mm against the test microorganisms. Compound 8 showed no activity against Gram-negative bacteria, Escherichia coli and Pseudomonas aeruginosa.

Minimum inhibitory concentration assay was performed in order to get more insight into the antimicrobial activity of compound 8 as a new antimicrobial pyrimidinone derivative. Compound 8 displayed antimicrobial activity towards the tested bacteria and pathogenic fungi with MIC values in the range of 0.0619–0.1859 μmol/mL (Table 5).

4. Conclusions

6-(1,3-Benzodioxol-5-ylmethyl)-5-ethyl-2-{[2-(morpholin-4-yl)ethyl]sulfanyl}pyrimidin-4(3H)-one (8) was synthesized as a novel antimicrobial pyrimidinone derivative bearing 1,3-benzodioxole moiety. The 3D structure of the target compound 8 and its S-alkylation were confirmed via single crystal X-ray crystallography. The antimicrobial evaluation of the title compound 8 revealed its dual antibacterial (MIC value = 0.0619 towards S. aureus) and antifungal activities (MIC value = 0.1859 towards both C. albicans and A. niger).

Additional Materials

Crystallographic data of compound 8 have been deposited with the Cambridge Crystallographic Data Center (supplementary publication number CCDC-991136). Copies of the data may be obtained free of charge from the Director, CCDC, 12 Union Road, Cambridge, CB2 1EZ, UK (deposit@ccdc.cam.ac.uk).

Conflict of Interests

The authors have declared that there is no conflict of interests.

Acknowledgment

The authors would like to extend their sincere appreciation to the Deanship of Scientific Research at King Saud University for its funding of this research though the Research Group Project no. RGP-VPP-196.

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