INTRODUCTION:

The Heterocyclic compounds comprising nitrogen, oxygen and sulphur heteroatom shows the number of functions in different fields because of their definite features. The nitrogen containing heterocyclic compounds integrated remarkably involved in the past studies of heterocyclic chemistry with the growth of synthetic route. Chalcones and cyclic imides perform a significant role in the heterocyclic synthesis containing nitrogen groups. Cyclic imides^[1] like succinimides^[2,3], maleimides^[4], glutarimide^[5] and phthalimides^[6] showed the defensive antibacterial^[7], antifungal activities. The substituted heterocyclic imides were developed from cyclic anhydrides^[8], form amide^[9], trifluoroacetylation^[10] and bis-heterocyclic analogs^[11] by microwave synthesis. Bis-chalcones are are prepared by the condensation^[12] of the substituted ketones and aldehyde groups^[13]. The chalcones are synthesized by utilizing a number of synthetic routes like solid phase Claisen-Schemdit, mixed-aldol condensation, acid catalyst^[14], coupling reaction^[15] and microwave assisted synthesis^[16].

The amino-pyrimidine derivative has been developed by the intermediary chalcones with nitrate, chlorides and carbonates of guanidine cyclized by ring expansion of N-C-N groups. A variety of novel amino-pyrimidines^[17], imido pyrimidines^[18], pyrido-pyrimidines^[19] and substituted pyrimidines from the chalcone intermediates^[20,21] revealed the significant antibacterial and antifungal activities^[22]. Several amino-pyrimidine derivatives exhibited the significant, antimicrobial^[23], antioxidant^[24], anticancer^[25], anti-plasmodial^[26], anti-proliferative leukemia^[27], breast carcinoma cells^[28], analgesic^[29] and apoptosis inducing activities^[30]. The synthesis of pyrimidine and amino-pyrimidine derivatives are prepared by conventional method using Claisen-Schmidt condensation liaising by chalcones^[31], cyclization of guanidine hydrochloride^[32], guanidine^[33]and guanidine nitrate^[34]. By eco-friendly point of view, pyrimidines and fused pyrimidines^[35], form amides^[36] and hydroxylamine^[37] by using clean, solvent free microwave synthesis.

EXPERIMENTAL SECTION:

Material methods

Melting points of all the synthesized compounds were recorded in an open glass capillaries and were uncorrected. IR spectra in (KBr pallets) were chronicled on Shimadzu FTIR-8400S and ATR Brucker alpha FT-IR spectrophotometer. ¹H NMR spectra were recorded on 400 MHz and 500.13 MHz by Brucker spectrophotometer. The reaction was monitored by thin layer chromatography was carried out through pre-coated silica gel aluminium plates with mixture of diethyl ether and ethyl acetate 7:3 proportion or benzene. All the compounds 3a-j was synthesized in the microwave oven in hours from bis-chalcones, guanidine nitrate, neutral alumina (Al2O3) and ethanol.

General procedure for the synthesis of amino-pyrimidines (3a-j):

To accomplish the work amino-pyrimidine (3a-j) derivatives were synthesized by the mixture of 0.2 moles of previously prepared bis-chalcones (1a-j) and 0.4 moles of guanidine nitrate in 2 gm of neutral Al2O3 under microwave supported solvent free condition on 640W power for 4-7 minutes. The synthesized compounds were recovered by ethyl acetate and recrystallized by ethanol (Scheme-I).

9-(1-phenyl)-4,5-(2","'-methoxyphenol)-9H-1,3,6,8,9-penta-azo-fluorene-2,7-diamine (3a)

Compound appearance: Yellowish Granular Crystals; M. F.: C₂₈H₂₃N₇O₄; Yield: 92.30%; M. Wt.: 521.53; M. P. (ºC): 179-181 ºC; Elt. Anal: C, 64.83; H, 4.73; N, 19.10; FTIR (ATR): -NH₂(2-Peaks): 3194 cm^-1 and 3315 cm^-1, aromatic ring (3-Peaks): 1500 cm^-1, 1578 cm^-1 and 1662 cm^-1, Ar-OCH₃: 1182 cm^-1, Ar-OH: 3400 cm^-1; ¹H NMR (300.06 MHz, DMSO-d⁶, δ ppm): 7.02-7.41 (m, 8H, Ar-H), 3.56 (s, 3H, OCH3), 9.73 (s, 2H, NH₂), 10.25 (s, 1H, -OH)

9-(4'-bromophenyl)-4,5-(2","'-methoxyphenol)-9H-1,3,6,8,9-penta-azo-fluorene-2,7-diamine (3b)

Compound appearance: Whitish Granular Crystals; M. F.: C₂₈H₂₃N₇O₄; Yield: 79.66%; M. Wt.: 600.42; M. P. (ºC): 117-119 ºC; Elt. Anal: C, 56.38; H, 3.84; N, 16.59; FTIR (KBr): -NH₂ (2-Peaks): 3255 cm^-1 and 3362 cm^-1, aromatic ring (3-Peaks): 1512 cm^-1, 1591 cm^-1 and 1669 cm-1, Ar-OCH₃: 1170 cm^-1, Ar-OH: 3524 cm^-1, Ar-Br: 1070 cm^-1

9-(4'-chlorophenyl)-4,5-(2","'-methoxyphenol)-9H-1,3,6,8,9-penta-azo-fluorene-2,7-diamine (3c)

Compound appearance: Whitish Pink Granular Crystals; M. F.: C₂₈H₂₃N₇O₄; Yield: 76.81%; M. Wt.: 555.97; M. P. (ºC): 104-106 ºC; Elt. Anal: C, 60.83; H, 4.25; N, 17.58; FTIR (KBr): -NH₂ (2-Peaks): 3301 cm^-1 and 3363 cm^-1, aromatic ring (3-Peaks): 1514 cm^-1, 1541 cm^-1 and 1594 cm^-1, Ar-OCH3: 1166 cm^-1, Ar-OH: 3595 cm^-1, Ar-Cl: 1094 cm^-1

9-(p-tolyl)-4,5-(2","'-methoxyphenol)-9H-1,3,6,8,9-penta-azo-fluorene-2,7-diamine (3d)

Compound appearance: Whitish Needle Shaped Crystals; M. F.: C₂₈H₂₃N₇O₄; Yield: 80.45%; M. Wt.: 535.55; M. P. (ºC): 108-110 ºC; Elt. Anal: C, 65.37; H, 4.94; N, 18.62; FTIR (KBr): -NH₂ (2-Peaks): 3304 cm^-1 and 3417 cm^-1, aromatic ring (3-Peaks): 1517 cm^-1, 1594 cm^-1 and 1673 cm^-1, Ar-OCH₃: 1182 cm^-1, Ar-OH: 3585 cm^-1

9-(4'-methoxyphenyl)-4,5-(2","'-methoxyphenol)-9H-1,3,6,8,9-penta-azo-fluorene-2,7-diamine (3e)

Compound appearance: Whitish Yellow Needle Shaped Crystals; M. F.: C₂₉H₂₅N₇O₅; Yield: 67.88%; M. Wt.: 551.55; M. P. (ºC): 105-107 ºC; Elt. Anal: C, 63.48; H, 4.75; N, 17.96; FTIR (KBr): -NH₂ (2-Peaks): 3300 cm^-1 and 3416 cm^-1, aromatic ring (3-Peaks): 1512 cm^-1, 1592 cm^-1and 1668 cm^-1, Ar-OCH₃: 1267 cm^-1, Ar-OH: 3612 cm^-1

9-(4'-fluorophenyl)-4,5-(2","'-methoxyphenol)-9H-1,3,6,8,9-penta-azo-fluorene-2,7-diamine (3f)

Compound appearance: Yellow Flakes; M. F.: C₂₈H₂₂FN₇O₄; Yield: 81.34%; M Wt.: 539.52; M. P. (ºC): 114-116 ºC; Elt. Anal: C, 62.65; H, 4.39; N, 18.47; FTIR (KBr): -NH₂(2-cm^-1, Ar-OCH₃: 1267 cm^-1, Ar-OH: 3612 cm^-1, Ar-F: 1173 cm^-1; ¹H NMR (500.03 MHz, DMSO-d⁶, δ ppm): 6.96-7.58 (m, 5H, Ar-H), 3.81 (s, 3H, OCH₃), 9.78 (s, 2H, NH₂), 10.27 (s, 1H, -OH).

9-(4'-nitrophenyl)-4,5-(2","'-methoxyphenol)-9H-1,3,6,8,9-penta-azo-fluorene-2,7-diamine (3g)

Compound appearance: Yellow Granular Crystals; M. F.: C₂₈H₂₂N₈O₆; Yield: 55.83%; M. Wt.: 566.52; M. P. (ºC): 113-115 ºC; Elt. Anal: C, 59.84; H, 4.35; N, 19.99; FTIR (KBr): -NH₂ (2-Peaks): 3223 cm^-1 and 3279 cm^-1, aromatic ring (3-Peaks): 1573 cm^-1, 1596 cm^-1 and 1676 cm^-1, Ar-OCH₃: 1205 cm^-1, Ar-OH: 3609 cm^-1, Ar-NO₂: 1503 cm^-1.

9-(naphthalen-4-yl)-4,5-(2","'-methoxyphenol)-9H-1,3,6,8,9-penta-azo-fluorene-2,7-diamine (3h)

Compound appearance: Pinkish Flakes; M. F.: C₃₂H₂₅N₇O₄; Yield: 82.11%; M. P. (ºC): 100-102 ºC; M. Wt.: 571.59; Elt. Anal: C, 67.56; H, 4.76; N, 17.83; FTIR (KBr): -NH₂ (2-Peaks): 3271 cm^-1 and 3264 cm^-1, aromatic ring (3-Peaks): 1463 cm^-1, 1512 cm^-1, 1591 cm^-1, 1671 cm^-1 and 1709 cm^-1, Ar-OCH₃: 1178 cm^-1, Ar-OH: 3612 cm^-1

9-(3'-chloro-4'-fluorophenyl)-4,5-(2","'-methoxyphenol)-9H-1,3,6,8,9-penta-azo-fluorene-2,7-diamine (3i)

Compound appearance: White Granular Crystals; M. F.: C₂₈H₂₁ClFN₇O₄; Yield: 83.50%; M. Wt.: 573.96; M. P. (ºC): 117-119 ºC; Elt. Anal: C, 58.88; H, 3.97; N, 17.39; FTIR (KBr): -NH₂ (2-Peaks): 3079 cm^-1 and 3206 cm^-1, aromatic ring (3-Peaks): 1511 cm^-1, 1589 cm^-1 and 1667 cm^-1, Ar-OCH3: 1266 cm^-1, Ar-OH: 3612 cm^-1, Ar-F: 1172 cm^-1, Ar-Cl: 1027 cm-1; ¹H NMR (500.03 MHz, DMSO-d⁶, δ ppm): 6.87-7.94 (m, 6H, Ar-H), 3.39 (s, 3H, OCH₃), 8.58 (s, 2H, NH₂), 9.72 (s, ¹H, -OH)

9-(2',4',5'-trichlorophenyl)-4,5-(2","'-methoxyphenol)-9H-1,3,6,8,9-penta-azo-fluorene-2,7-diamine (3j)

Compound appearance: White Needle Shaped Crystals; M. F.: C₂₈H₂₀C_l3N₇O₄; Yield: 73.71%; M. Wt.: 624.86; M. P. (ºC): 149-151 ºC; Elt. Anal: C, 53.89; H, 3.67; N, 15.89; FTIR (KBr): -NH₂ (2-Peaks): 3169 cm^-1and 3291 cm^-1, aromatic ring (3-Peaks): 1512 cm^-1, 1576 cm^-1and 1667 cm^-1, Ar-OCH₃: 1212 cmcm^-1, Ar-OH: 3601 cm^-1, Ar-2,4,5Cl: 1078 cm^-1

RESULT AND DISCUSSION;

Chemistry

The starting compounds of bis-chalcones 1a-j were prepared by the reaction of substituted N-phenyl succinimides using di-substituted aromatic aldehyde i.e. vanillin. Thesolvent free condition. Formation of amino-pyrimidines was confirmed by IR, ¹HNMR and elemental analysis.

Antimicrobial activities (3a-j)

All the synthesized compounds 3a-j were Screened for their antibacterial activity against gram positive bacteria Bacillus subtilis (MCMB-310) and gram negative bacteria Escherichia coli (MCMB-301) using DMF solvent. And antifungal activities against Candida albicans (NCIM-3471) and Aspergillus niger (NCIM- 545) strains using DMSO solvent. Moderate to good antibacterial activities shown by Bacillus subtilis and Escherichia coli and similarly the compound these shown significantly synergistic and superior antifungal activities against Candida albicans and Aspergillus niger strains. Ampicillin used for antibacterial and Amphotericin-B used for antifungal activities as a standard drugs as shown in the graph-1. The calculated readings are put into a table -1.

Table-1: Antimicrobial activities of Bis-chalcones

Compd Code	Zone diameter calculated in mm and tabulated by (Mean±S.D.)
	*Bacillus subtilis*	*Escherichia coli*	*Candida albicans*	*Aspergillus niger*
	100 μg/ml	100 μg/ml	100 μg/ml	100 μg/ml
3a	6.66±0.57 **	7.33±0.57 **	11.48 ± 0.10 **	--
3b	7±1 **	7±0 **	12.85 ± 0.24 **	--
3c	8.33±1.15 **	8.66±0.57 **	18.16 ± 0.24 **	15.45 ± 0.18 **
3d	6.33±0.57 **	8.66±0.57 **	20.00 ± 0.27 **	13.64 ± 0.22 **
3e	6.66±1.15 **	7.33±0.57 **	15.16 ± 0.27 **	14.63 ± 0.22 **
3f	8.33±2.08 **	8±0 **	20.64 ± 0.17 **	13.37 ± 0.24 **
3g	7±1 **	8.33±0.57 **	14.82 ± 0.76 **	16.12 ± 0.32 **
3h	7±1 **	7.33±0.57 **	20.29 ± 0.53 **	17.34 ± 0.12 **
3i	7.33±0.57 **	7.33±0.57 **	22.50 ± 0.40 **	20.54 ± 0.21 **
3j	8.66±0.57 **	5±4.35 **	15.74 ± 0.13 **	16.54 ± 0.25 **
Ctrl	0.0±0.0	0.0±0.0	0.0 ± 0.0	0.0 ± 0.0
Std	18.33±0.57	18.33±0.57	12.40 ± 0.43	10.45 ± 0.11
Keynote: Zone of inhibition measured in mm (Mean±S.D.) (N=3) ('--' means no zone)

Graph 1: Antimicrobial activities of 3a-j (Mean ± SD)

Statistical Analysis

All the results of the synthesized compound series 3a-j were carried out by the triplicate format N=3 with Mean ± SD. The statistical tests were performed by using Graph Pad in Stat software. The statistical significance was brought by one way ANOVA and confirmed by Dunnett multiple comparisons test performed the standard drugs against synthesized compounds. P value < 0.05 was considered as statistically significant remarked by *p<0.05, **p<0.01, ***p<0.001 compared to standard groups.

CONCLUSION:

A method for synthesis of amino-pyrimidines 3a-j has been developed in good yield obtained by the treatment of substituted bis-chalcones and guanidine nitrate with neutral Al₂O₃. All these compounds were characterized by their spectral analysis. Most of the compound showed moderate activity against Bacillus subtilis and Escherichia coli. Correspondingly these compounds exhibited significant as well as synergistic activities against Candida albicans and Aspergillus niger fungal strains. All the synthesized compounds may be used for preparation of various heterocyclic systems by using microwave method.

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Received on 05.12.2018 Modified on 25.12.2018

Research J. Pharm. and Tech 2019; 12(1): 167-171.

DOI: 10.5958/0974-360X.2019.00031.3