Synthesis and biological evaluation of novel series of aminopyrimidine derivatives as urease inhibitors and antimicrobial agents

Article abstract of DOI:10.3109/14756366.2012.740477

A novel series of carbazole substituted aminopyrimidines (5a-p) were synthesized and screened for their in vitro urease inhibition and antimicrobial activity. Among the compounds, 4-(2,4-dichlorophenyl)-6-(9-methyl-9H-carbazol-3- yl)-pyrimidin-2-amine (5i) was found to be the most potent showing urease inhibitory activity with an IC50 value 19.4±0.43 μM. Compounds 5c, 5g, 5j and 5o showed good activity against all selected bacterial strains and compounds 5b, 5c, 5m and 5o showed good activity against selected fungal strains. All the compounds were subjected for ADME predictions by computational method.

Full text of DOI:10.3109/14756366.2012.740477

Journal of Enzyme Inhibition and Medicinal Chemistry, 2012; Early Online: 1–8
© 2012 Informa UK, Ltd.
ISSN 1475-6366 print/ISSN 1475-6374 online
DOI: 10.3109/14756366.2012.740477
RESEARCH ARTICLE
Synthesis and biological evaluation of novel series of
aminopyrimidine derivatives as urease inhibitors and
antimicrobial agents
Laxman K. Adsul1, Babasaheb P. Bandgar1, Hemant V. Chavan1, Shivkumar S. Jalde1, Valmik D.
Dhakane1, and Amol L. Shirfule2
1Medicinal Chemistry Research Laboratory, School of Chemical Sciences, Solapur University, Solapur, Maharashtra,
India and 2Food and Drug Toxicology Research Centre, National Institute of Nutrition, Hyderabad, Andhra Pradesh,
India
Abstract
A novel series of carbazole substituted aminopyrimidines (5a-p) were synthesized and screened for their in vitro
urease inhibition and antimicrobial activity. Among the compounds, 4-(2,4-dichlorophenyl)-6-(9-methyl-9H-
carbazol-3-yl)-pyrimidin-2-amine (5i) was found to be the most potent showing urease inhibitory activity with an
IC50 value 19.4 0.43 µM. Compounds 5c, 5g, 5j and 5o showed good activity against all selected bacterial strains
and compounds 5b, 5c, 5m and 5o showed good activity against selected fungal strains. All the compounds were
subjected for ADME predictions by computational method.
Keywords: Carbazole, aminopyrimidines, urease inhibition, antimicrobial activity
Introduction
its derivatives, and inorganic metal salts, etc9–11. Various
heterocyclic compounds were reported as urease
inhibitors by several research groups. Khan et al. have
synthesized biscoumarins which were shown as urease
inhibitors8. Akhtar et al. have synthesized 3-substituted-
Urease inhibitors are considered as new targets for antiul-
cer drugs1. e activity of bacterial urease has been shown
to be important virulence factor in the development of
many harmful clinical conditions for human and animal
health as well as agriculture2. Bacterial ureases have been
reported to be involved in the formation of infectious
stones and development of peptic ulcers and stomach
cancer3. Ureases also contribute to the development of
urolithiasis, pyelonephritis, hepatic encephalopathy,
hepatic coma, and urinary catheter encrustation4. In the
near past, a number of compounds have been proposed
as urease inhibitors to reduce environmental problems
and enhance the uptake of urea nitrogen by plants5–8. e
treatment of infections caused by urease producing bac-
teria may also be possible by urease inhibition. In recent
years, a variety of urease inhibitors have been studied,
including acetohydroxamic acid, flurofamide, omepra-
zole, lansoprazole, rabaprazole, 1,4-benzoquinone and
4-amino-5-thioxo-1H-4H-1,2,4-triazoles
derivatives
showing their urease inhibition activity12. Rauf et al. have
reported the synthesis and urease inhibition activity of
barbituric and thiobarbituric acid derivatives13. More
recently, Khan et al. also reported arylidene barbiturates
as urease inhibitors14. Amines derived from deoxybenzo-
ins are also reported as urease inhibitors15.
e pyrimidine entity is one of the most prominent
structures found in nucleic acid chemistry. Pyrimidine
derivatives including uracil, thymine, cytosine, adenine,
and guanine are fundamental building blocks for
deoxyribonucleic acid (DNA) and ribonucleic acid (RNA).
Vitamin B1 (thiamine) is a well-known example of a
naturally occurring pyrimidine that is encountered in our
Address for Correspondence: Babasaheb P. Bandgar, Medicinal Chemistry Research Laboratory, School of Chemical Sciences, Solapur
University, Solapur 413 255, Maharashtra, India. Tel/Fax: 0091-217-2351300. E-mail: bandgar_bp@yahoo.com
(Received 12 July 2012; revised 13 September 2012; accepted 09 October 2012)
1

2
L. K. Adsul et al.
daily lives16,17. Pyrimidine derivatives form a component
in a number of useful drugs and are associated with many
biological, pharmaceutical and therapeutical activities18.
Condensed pyrimidine derivatives have been reported as
antimicrobial19, analgesic20, antiviral, anti-inflammatory21,
anti-HIV22, antitubercular23, antitumour24, antineoplastic25,
antimalarial26, diuretic27 and antiplatelet agents28.
Like pyrimidines, carbazole compounds also exhibits
diverse biological properties. Carbazole and its deriva-
tives are the important class of heterocyclic compounds
endowed with various pharmacological activities such as
anticancer, antimicrobial, antiviral, anti-inflammatory
and antioxidant activity29. Carbazole scaffold is pres-
ent in many drugs such as carvedilol and carprofen
(Figure 1). Carvedilol, an antihypertensive drug, acts as
a nonspecific β-adrengic antagonist30. Carprofen, a non-
steroidal anti-inflammatory drug (NSAID), is a selective
COX-2 inhibitor31. More recently, a series of phenylcarba-
zole (Figure 1) molecules have been reported as antitu-
mour agent32, carbazole sulphonamides (Figure 1) are a
novel class of antimitotic agents against solid tumours33.
Syutkin et al.34 have reported carbazole containing chal-
cones and pyrimidines as light emitting diodes.
Itwasenvisagedthatthesetwoactivepharmacophores,if
linked together would generate novel molecular templates
which are likely to exhibit interesting biological properties.
e above-cited applications pyrimidines and barbiturates
prompted us to synthesize a series of carbazole contain-
ing pyrimidine derivatives. Owing to this importance and
in continuation of our work on synthesis of biologically
active compounds35–37, now we wish to describe the syn-
thesis of carbazole containing aminopyrimidine deriva-
tives (Scheme 1). e newly synthesized compounds were
screened for their in-vitro urease inhibition and antimicro-
bial activity. We also subjected all synthesized compounds
for ADME predictions by computational method.
iodide gave 9-methyl carbazole (2), which on Vilsmeier-
Haack formylation gave 3-formyl-9-methylcarbazole
(3). e 3-formyl-9-methylcarbazole (3) on Claisen-
Schimdt condensation with various aromatic acetophe-
nones and heteroaromatic acetophenones in aqueous
sodium hydroxide afforded compounds 4a-p37. is on
further treatment with guanidine hydrochloride in pres-
ence of NaH in DMF gave the target compounds 5a-p.
Compounds have been characterized by IR, 1H NMR
1
and mass spectroscopy. In the H NMR spectra of these
compounds, the pyrimidine ring proton was observed
downfield as a singlet at δ ~7.80 and NH2 proton appears
at δ ~6.65 as a singlet.
Biological evaluation
e urease inhibition activity was carried out according
to the literature protocol38 using thiourea as the stan-
dard inhibitor having an IC50 value of 21.0 0.14 µM and
the results are presented in Table 1. Out of 16 screened
compounds, compounds 5c, 5f, 5g, 5i, 5j, and 5k were
found to be active against urease. Compound 5i proved
to be the most potent showing urease inhibitory activ-
ity with an IC50 value 19.4 0.23 µM as compared to the
standard thiourea. e compounds 5j (IC50 = 23.2 0.58)
and 5k (IC50 = 28.1 0.59) exhibited good activity with
an IC50 value as compared to that of standard thio-
urea. Compound 5c with 3,4-dimethoxy substitu-
ent was found to be good inhibitor with an IC50 value
of 30.7 0.94. Compounds 5f and 5g were displayed
moderate urease inhibitory activities with an IC50 value
40.2 0.31 and 45.2 0.63, respectively. From these
results of urease inhibitory activities by pyrimidine
derivatives, following tentative results can be drawn; it
appears that disubstituted halogen compounds were
found to inhibit urease enzyme. Dichloro substituted
compounds were showing promising urease inhibi-
tory activity than difluoro and dimethoxy substituted
compounds. Whereas, monosubstituted compound
with electron donating and withdrawing substituent
are weak inhibitors of urease. Heteroaryl substituted
pyrimidine derivatives (compounds 5n and 5o) are also
weak inhibitors of urease.
Results and discussion
Chemistry
e target compounds were synthesized as shown in
Scheme 1. Carbazole (1) on methylation with methyl
e synthesized pyrimidine derivatives were tested for
their in vitro antimicrobial against five bacterial and two
fungal species using disk diffusion method. Tetracycline
and nystatin were used as standards against bacteria
and fungi, respectively. e antibacterial activity against
two Gram positive bacteria i.e. Bacillus subtilis (BS) and
Staphylococcus aureus (SA), and three Gram negative
bacteria, i.e. Escherichia coli (EC), Shigella flexenari (SF)
and Salmonella typhi (ST) was tested at a sample con-
centration of 1 mg/mL in DMSO. e antibacterial activ-
ity data is tabulated in Table 2. e result indicated that
among the tested compounds, compounds 5c, 5g, 5j
and 5o showed good activity against all selected bacte-
rial strains at concentration of 1 mg/mL as compared to
standard drug tetracycline. Compound 5o showed com-
parable activity as that of standard, against B. subtilis
Figure 1. Pharmacologically active carbazole derivatives.
Journal of Enzyme Inhibition and Medicinal Chemistry

Novel series of aminopyrimidine derivatives
3
Scheme 1. Reagents and conditions: (i) CH3I, NaH, DMF, rt, 3 h; (ii) DMF, POCl3, 80°C, 3 h; (iii) Substituted acetophenones, NaOH, Ethanol,
rt, 24 h; (iv) Guanidine.HCl, DMF, NaH, 110°C, 10 h.
result indicated that among the tested compounds, com-
pounds 5b, 5c, 5m and 5o showed good activity against
Table 1. Urease inhibition activity.
Compound
IC50 SEM (µM)*
>100
selected fungal strains at concentration of 1mg/mL as
compared to standard drug nystatin. Compounds 5m
and 5o showed comparable activity as that of standard,
against C. albicans (CA) and A. niger (AN). e remaining
compounds showed moderate activity against C. albicans
(CA). While, all remaining compounds showed less activ-
ity against A. niger (AN).
5a
5b
5c
5d
5e
5f
5g
5h
5i
5j
95.4 0.52
30.7 0.94
73.7 0.45
>100
40.2 0.31
45.2 0.63
>100
e bioavailability of the target compounds was
assessed using ADME (absorption, distribution, metabo-
lism, and excretion) prediction methods. In particular,
we calculated the compliance of compounds to the
Lipinski’s rule of five39,40. Predictions of ADME proper-
ties for the studied compounds are shown in Table 3. e
results showed that all the tested compounds comply
with the rules. Compounds 5h, 5i, 5j, 5k and 5l have c
Log P value more than 5 however, these compounds have
molecular weight, hydrogen bond donor, hydrogen bond
acceptor within the range of Lipinski rule of five and these
compounds have low polar surface area (PSA) and molar
volume (MV). Molecular polar surface area (PSA) is a
very useful parameter for the prediction of drug transport
properties. All the tested compounds have maximum
absorption as they are having low PSA and MV. Drug-
likeness score (a combined effect of physico-chemical
properties, pharmacokinetics and pharmacodynamics
of a compound and is represented by a numerical value)
was computed by MolSoft (MolSoft 2007) software for
19.4 0.43
23.2 0.58
28.1 0.59
95.1 0.43
87.2 0.60
>100
5k
5l
5m
5n
5o
5p
iourea
>100
83.7 0.94
21.0 0.14
*IC50 values represent mean SD from three different experiments.
(BS), S. aureus (SA) and S. flexenari (SF). e remaining
compounds showed moderate activity against all of the
tested bacterial strains except S. typhi (ST).
e antifungal activity of pyrimidine derivatives
against two fungal species, i.e. Candida albicans (CA) and
Aspergillus niger (AN) was tested at a sample concentra-
tion of 1mg/mL in DMSO using nystatin as standard.
e antifungal activity data are tabulated in Table 2. e
© 2012 Informa UK, Ltd.

4
L. K. Adsul et al.
the 16 molecules under study. Maximum drug-likeness
score was found to be 0.85 for the compound 5b. Hence,
these compounds do not pose absorption and perme-
ation difficulty.
the most potent showing urease inhibitory activity with
an IC50 value 19.4 0.43 µM. Compounds 5c, 5g, 5j and
5o showed good activity against all selected bacterial
strains and compounds 5b, 5c, 5m and 5o showed good
activity against selected fungal strains. ADME prediction
revealed that the compounds could be considered as
good candidates for drug development.
Conclusion
A new series of carbazole substituted pyrimidine deriva-
tives were synthesized and screened for their in-vitro ure-
ase inhibition and antimicrobial activity. Compounds 5c,
5f, 5g, 5i, 5j, and 5k were found to be active against ure-
ase. Compound, 4-(2,4-dichlorophenyl)-6-(9-methyl-
9H-carbazol-3-yl)-pyrimidin-2-amine 5i was found to be
Experimental section
General
All the reagents and solvents used were of analytical
grade and were used as supplied unless otherwise stated.
Table 2. Antimicrobial activity of pyrimidine derivatives.
Antibacterial activity
SA
Antifungal activity
Compound
EC
–
BS
–
SF
–
ST
–
CA
9
AN
8
5a
5b
5c
5d
5e
5f
5g
5h
14
11
14
10
12
11
16
8
–
11
13
10
12
16
12
9
13
12
8
–
14
12
9
12
12
–
9
9
–
–
–
11
10
13
9
–
12
13
14
8
9
–
–
10
8
8
–
–
–
8
5i
5j
5k
5l
5m
5n
5o
5p
–
12
13
11
10
11
9
13
14
12
13
12
10
18
15
20
–
14
12
11
10
11
12
18
14
16
–
8
–
12
9
10
9
9
–
9
–
10
–
8
–
–
–
–
14
9
10
–
8
–
10
9
18
12
20
–
–
15
10
–
12
8
–
Tetracycline
Nystatin
16
–
16
–
–
18
16
Concentration: 1 mg/mL in DMSO; Size of well 6 mm in diameter.
(–): Compounds showing no inhibition.
EC, E. coli; BS, B. subtilis; SA, S. aureus; SF, S. flexenari; ST, S. typhi; CA, C. albicans; AN, A. niger.
Table 3. In silico pharmacological parameters for bioavailability.
Compounds
MW
350
380
410
440
368
386
386
384
418
418
418
428
395
356
351
365
c log p
4.69
4.75
4.31
4.15
4.81
4.45
4.96
5.38
5.85
5.99
5.70
5.59
4.59
4.40
3.43
3.86
HBA
2
HBD
2
PSA
MV
Drug likeness score
−0.17
0.85
5a
5b
5c
5d
5e
5f
5g
5h
5i
5j
5k
5l
5m
5n
5o
5p
42.54
50.92
58.63
66.85
42.54
42.54
42.54
42.54
42.54
42.54
42.54
42.54
85.18
43.56
51.86
63.62
332.76
364.61
396.21
425.79
338.68
343.60
343.99
349.96
365.33
365.28
365.33
354.62
356.53
327.64
327.04
340.34
3
2
4
2
0.47
5
2
−0.36
−0.41
−0.26
−0.71
−0.21
0.03
2
2
2
2
2
2
2
2
2
2
2
2
−0.43
−0.35
−0.72
−0.51
0.52
2
2
2
2
3
2
3
2
3
2
−0.26
−0.85
2
4
HBA, hydrogen bond acceptors; HBD, hydrogen bond donors; MV, molecular volume; PSA, polar surface area.
Journal of Enzyme Inhibition and Medicinal Chemistry

Novel series of aminopyrimidine derivatives
5
Melting points were recorded in open capillaries with
electrical melting point apparatus and were uncorrected.
IR spectra (KBr disks) were recorded using a Perkin–
Elmer 237 spectrophotometer. 1H NMR (200 MHz) spec-
tra were recorded on a Bruker Avance spectrometer using
DMSO-d6 as solvent. Mass spectra were recorded on a
Shimadzu LCMS-QP 1000 EX. in layer chromatogra-
phy (TLC) was performed on silica gel-coated plates for
monitoring the reactions.
4-(9-methyl-9H-carbazol-3-yl)-6-(2,4,6-trimethoxyphenyl)-
pyrimidin-2-amine (5d)
Yield: 86%; MP: 208–211°C; IR (KBr, cm−1): 3368, 3320,
1
3214, 3019, 1641, 1559, 1372, 1220, 1149, 930; H NMR
(200 MHz, DMSO-d6, δ in ppm): 3.80 (s, 3H, OCH3), 3.85
(s, 6H, OCH3), 4.01 (s, 3H, NCH3), 6.35 (s, 2H, ArH), 6.66
(s, 2H, NH2), 7.38 (t, 1H, J = 8 Hz, ArH), 7.58 (t, 1H, J = 8 Hz,
ArH), 7.61–7.70 (m, 1H, ArH), 7.90 (s, 1H, ArH), 8.00–8.08
(m, 2H, ArH), 8.31 (d, 1H, J = 8 Hz, ArH), 9.24 (s, 1H, ArH);
MS: m/e 441 (M + 1).
General procedure for the synthesis of 4-(9-methyl-9H-
carbazol-3-yl)-6-phenylpyrimidin-2-amine (5a-p)
4-(4-fluorophenyl)-6-(9-methyl-9H-carbazol-3-yl)-pyrimidin-
2-amine (5e)
Chalcones 4a-p (1.0 mmol) was dissolved in DMF
(15 mL) and guanidine hydrochloride (2 mmol) was
added to it. To this reaction mixture, NaH (4.0 mmol) was
added and the reaction mixture was heated up to 110°C
for 10 h. After completion of reaction (TLC), the reac-
tion mixture was cooled to room temperature and water
(15 mL) was added to it, solid product started to separate
out. e solid was filtered and purified by column chro-
matography using chloroform as eluent to afford target
compounds 5a-p.
Yield: 92%; MP: 230–232°C; IR (KBr, cm−1): 3417, 3330,
3296, 3020, 1596, 1566, 1443, 1369, 1212, 1155, 1045, 932,
663; 1H NMR (200 MHz, DMSO-d6, δ in ppm): 3.85 (s, 3H,
NCH3), 6.66 (s, 2H, NH2), 7.23–7.58 (m, 6H, ArH), 7.80 (s,
1H, ArH), 8.23–8.28 (m, 4H, ArH), 9.12 (s, 1H, ArH); MS:
m/e 369 (M + 1).
4-(2,4-difluorophenyl)-6-(9-methyl-9H-carbazol-3-yl)-
pyrimidin-2-amine (5f)
Yield: 87%; MP: 112–116°C; IR (KBr, cm−1): 3417, 3330,
3296, 3020, 1596, 1566, 1443, 1369, 1212, 1155, 1140,
Spectral data for synthesized compounds
Synthesis of 4-(9-methyl-9H-carbazol-3-yl)-6-
phenylpyrimidin-2-amine (5a)
1
930; H NMR (200 MHz, DMSO-d6, δ in ppm): 3.85 (s,
3H, NCH3), 6.66 (s, 2H, NH2), 7.23 (t, 1H, J = 8 Hz, ArH),
7.48–7.58 (m, 3H, ArH), 7.61–7.73 (m, 2H, ArH), 7.80 (s,
1H, ArH), 8.23–8.28 (m, 2H, ArH), 8.40 (d, 1H, J = 8 Hz,
ArH), 9.08 (s, 1H, ArH); MS: m/e 387 (M + 1).
Yield: 90%; MP: 234–236°C; IR (KBr, cm−1): 3368, 3325,
1
3214, 3020, 1641, 1559, 1372, 1216, 1149; H NMR (200
MHz, DMSO-d6, δ in ppm): 3.82 (s, 3H, NCH3), 6.66 (s,
2H, NH2), 7.23 (t, 1H, J = 8 Hz, ArH), 7.48–7.58 (m, 4H,
ArH), 7.61–7.73 (m, 2H, ArH), 7.80 (s, 1H, ArH), 8.23–8.28
(m, 3H, ArH), 8.40 (d, 1H, J = 8 Hz, ArH), 9.07 (s, 1H, ArH);
MS: m/e 351 (M + 1).
4-(3,4-difluorophenyl)-6-(9-methyl-9H-carbazol-3-yl)-
pyrimidin-2-amine (5g)
Yield: 83%. MP: 190–192°C; IR (KBr, cm−1): 3417, 3330,
3296, 3020, 1596, 1566, 1443, 1369, 1212, 1155, 1140,
1
930; H NMR (200 MHz, DMSO-d6, δ in ppm): 3.85 (s,
4-(4-methoxyphenyl)-6-(9-methyl-9H-carbazol-3-yl)-
pyrimidin-2-amine (5b)
3H, NCH3), 6.66 (s, 2H, NH2), 7.23 (t, 1H, J = 8 Hz, ArH),
7.48–7.58 (m, 3H, ArH), 7.61–7.73 (m, 2H, ArH), 7.80 (s,
1H, ArH), 8.23–8.28 (m, 2H, ArH), 8.41 (d, 1H, J = 8 Hz,
ArH), 9.07 (s, 1H, ArH); MS: m/e 387 (M + 1).
Yield: 87%; MP: 210–212°C; IR (KBr, cm−1): 3368, 3320,
1
3214, 3019, 1641, 1559, 1372, 1220, 1149, 930; H NMR
(200 MHz, DMSO-d6, δ in ppm): 3.80 (s, 3H, OCH3), 3.87
(s, 3H, NCH3), 6.66 (s, 2H, NH2), 6.90 (d, 2H, J = 8 Hz,
ArH), 7.23 (t, 1H, J = 8 Hz, ArH), 7.48–7.58 (m, 2H, ArH),
7.61–7.73 (m, 2H, ArH), 7.80 (s, 1H, ArH), 8.23–8.28 (m,
2H, ArH), 8.40 (d, 1H, J = 8 Hz, ArH), 9.07 (s, 1H, ArH);
MS: m/e 381 (M + 1).
4-(4-chlorophenyl)-6-(9-methyl-9H-carbazol-3-yl)-pyrimidin-
2-amine (5h)
Yield: 93%. MP: 220–222°C; IR (KBr, cm−1): 3350, 3019,
1
2926, 2400, 1597, 1535, 1425, 1123, 1015, 928; H NMR
(200 MHz, DMSO-d6, δ in ppm): 3.87 (s, 3H, NCH3), 6.77
(s, 2H, NH2), 7.31 (t, 1H, J = 8 Hz, ArH), 7.48–7.58 (m, 5H,
ArH), 7.83 (s, 1H, ArH), 8.22–8.38 (m, 3H, ArH), 8.45 (d,
1H, J = 8 Hz, ArH), 9.12 (s, 1H, ArH); MS: m/e 385 (M + 1).
4-(3,4-dimethoxyphenyl)-6-(9-methyl-9H-carbazol-3-yl)-
pyrimidin-2-amine (5c)
Yield: 86%; MP: 218–221°C; IR (KBr, cm−1): 3368, 3320,
1
3214, 3019, 1641, 1559, 1372, 1220, 1149, 930; H NMR
4-(2,4-dichlorophenyl)-6-(9-methyl-9H-carbazol-3-yl)-
pyrimidin-2-amine (5i)
(200 MHz, DMSO-d6, δ in ppm): 3.82 (s, 3H, OCH3), 3.87
(s, 3H, OCH3), 4.00 (s, 3H, NCH3), 7.26 (d, 1H, J = 8 Hz,
ArH), 7.38 (t, 1H, J = 8 Hz, ArH), 7.58 (t, 1H, J = 8 Hz,
ArH), 7.70 (s, 1H, ArH), 7.81 (d, 1H, J = 8 Hz, ArH), 7.90
(s, 1H, ArH), 8.00–8.08 (m, 2H, ArH), 8.31 (d, 1H, J = 8 Hz,
ArH), 8.45–8.50 (m, 1H, ArH), 9.24 (s, 1H, ArH). MS: m/e
411 (M+1).
Yield: 82%; MP: 210–212°C; IR (KBr, cm−1): 3360, 3325,
1
3019, 2926, 2400, 1597, 1535, 1425, 1123, 1015, 930; H
NMR (200 MHz, DMSO-d6, δ in ppm): 3.87 (s, 3H, NCH3),
6.80 (s, 2H, NH2), 7.28 (t, 1H, J = 8 Hz, ArH), 7.48–7.58 (m,
5H, ArH), 7.83 (s, 1H, ArH), 8.25–8.38 (m, 3H, ArH), 9.02
(s, 1H, ArH); MS: m/e 419 (M+).
© 2012 Informa UK, Ltd.

6
L. K. Adsul et al.
4-(3,4-dichlorophenyl)-6-(9-methyl-9H-carbazol-3-yl)-
pyrimidin-2-amine (5j)
4-(3-aminophenyl)-6-(9-methyl-9H-carbazol-3-yl)-pyrimidin-
2-amine (5p)
Yield: 80%; MP: 184–187°C; IR (KBr, cm−1): 3360, 3325,
Yield: 70%; MP: 178–180°C; IR (KBr, cm−1): 3360, 3320,
3020, 2925, 1595, 1560, 1445, 1369, 1212, 1155, 1045,
930; 1H NMR (200 MHz, DMSO-d6, δ in ppm): 3.87
(s, 3H, NCH3), 6.40 (s, 2H, NH2), 6.79 (s, 2H, NH2), 7.23
(t, 1H, J = 8 Hz, ArH), 7.48–7.58 (m, 4H, ArH), 7.61–7.73
(m, 2H, ArH), 7.80 (s, 1H, ArH), 8.23–8.28 (m, 2H, ArH),
8.40 (d, 1H, J = 8 Hz, ArH), 9.09 (s, 1H, ArH); MS: m/e
366 (M + 1).
1
3019, 2926, 2400, 1597, 1535, 1425, 1123, 1015, 930; H
NMR (200 MHz, DMSO-d6, δ in ppm): 3.87 (s, 3H, NCH3),
6.80 (s, 2H, NH2), 7.32 (t, 1H, J = 8 Hz, ArH), 7.48–7.77 (m,
5H, ArH), 7.87 (s, 1H, ArH), 8.30 (d. 1H, J = 7.2 Hz, ArH),
8.37 (s, 1H, ArH), 8.48 (d, 1H, J = 8 Hz, ArH), 9.14 (s, 1H,
ArH); MS: m/e 419 (M+).
4-(2,5-dichlorophenyl)-6-(9-methyl-9H-carbazol-3-yl)-
pyrimidin-2-amine (5k)
Urease inhibition assay
Yield: 86%; MP: 210–212°C; IR (KBr, cm−1): 3360, 3325,
Mixture of 25 µL of urease enzyme (Jack bean) solution
containing 1 unit of enzyme (1 unit = 0.02 mg of enzyme)
was dissolved in phosphate buffer of pH 6.8, 55 µL of buf-
fer containing 100 mM urea, and 5 µL of test compounds
(0.5 mM concentration) was incubated at 30°C for 15 min
in 96-well plates. Urease activity was determined by
measuring ammonia production using the iodophenol
method38. Briefly, 45 µL each of phenol reagent and 70 µL
of alkali reagent were added to each well. e increase in
absorbance at 630 nm was measured after 50 min, using
a microplate reader (Molecular Device, CA, USA). All
reactions were performed in triplicate in a final volume
of 200 µL. e results (change in absorbance per min)
were processed using Soft-Max Pro software (Molecular
Device, CA, USA). iourea was used as the standard
inhibitor and percentage inhibitions were calculated as
follow: 100 − (ODtestwell/ODcontrol) × 100.
1
3019, 2926, 2400, 1597, 1535, 1425, 1123, 1015, 930; H
NMR (200 MHz, DMSO-d6, δ in ppm): 3.87 (s, 3H, NCH3),
6.80 (s, 2H, NH2), 7.28 (t, 1H, J = 8 Hz, ArH), 7.48–7.58 (m,
5H, ArH), 7.83 (s, 1H, ArH), 8.20–8.40 (m, 3H, ArH), 9.02
(s, 1H, ArH); MS: m/e 419 (M+).
4-(4-bromophenyl)-6-(9-methyl-9H-carbazol-3-yl)-pyrimidin-
2-amine (5l)
Yield: 82%; MP: 260–262°C; IR (KBr, cm−1): 3365, 3330,
3296, 3020, 1596, 1566, 1443, 1369, 1212, 1155, 1045, 932;
1H NMR (200 MHz, DMSO-d6, δ in ppm): 3.87 (s, 3H,
NCH3), 6.77 (s, 2H, NH2), 7.31 (t, 1H, J = 8 Hz, ArH),
7.48–7.58 (m, 5H, ArH), 7.81 (s, 1H, ArH), 8.25–8.37 (m,
3H, ArH), 8.40 (d, 1H, J = 8 Hz, ArH), 9.10 (s, 1H, ArH);
MS: m/e 431 (M + 2).
4-(9-methyl-9H-carbazol-3-yl)-6-(4-nitrophenyl)-pyrimidin-2-
amine (5m)
Antimicrobial activity
Yield: 94%; MP: 144–146°C; IR (KBr, cm−1): 3360, 3320,
Antimicrobial activities of all the synthesized compounds
were determined by disc diffusion method41. All the
bacterial and fungal species used were procured from
Institute of Microbial Type Culture Collection (IMTCC),
Chandigarh, India, and National Collection of Industrial
Microorganisms(NCIM),Pune,India,namely,Escherichia
coli, Bacillus subtilis, Staphylococcus aureus, Shigella flex-
enari, Salmonella typhi, Candida albicans, and Aspergillus
niger. e test compounds were dissolved in dimethyl
sulfoxide (DMSO) at a concentration of 1mg/mL. 20mL
of sterilized agar media was poured into each presteril-
ized Petri dish. Excess of suspension was decanted and
plates were dried by placing in an incubator at 37°C for
an hour. About 60 µL of 24-h-old culture suspension were
poured and neatly swabbed with the presterilized cotton
swabs. 6-mm diameter well was then punched carefully
using a sterile cork borer and 30 µL of test solutions were
added into each labelled well. Inoculated plates in dupli-
cate were then incubated at 37 0.5°C for antibacterial
activity for 24h and at 25 0.5°C for antifungal activity
for 48h. After incubation, the antimicrobial activity was
measured in terms of the zone of inhibition in mm.
1
3020, 1596, 1566, 1443, 1369, 1212, 1155, 1045, 932; H
NMR (200 MHz, DMSO-d6, δ in ppm): 3.85 (s, 3H, NCH3),
6.88 (s, 2H, NH2), 7.28 (t, 1H, J = 8 Hz, ArH), 7.48–7.74 (m,
4H, ArH), 8.02 (s, 1H, ArH), 8.23–8.65 (m, 5H, ArH), 9.11
(s, 1H, ArH); MS: m/e 396 (M + 1).
4-(9-methyl-9H-carbazol-3-yl)-6-(thiophen-2-yl)-pyrimidin-2-
amine (5n)
Yield: 80%; MP: 144–146°C; IR (KBr, cm−1): 3360, 3325,
1
3019, 1595, 1560, 1445, 1369, 1212, 1155, 1045, 928; H
NMR (200 MHz, DMSO-d6, δ in ppm): 3.87 (s, 3H, NCH3),
6.77 (s, 2H, NH2), 7.23 (t, 1H, J = 8 Hz, ArH), 7.35 (t, 1H,
J = 3.9 and 4.8 Hz, ArH), 7.48–7.60 (m, 3H, ArH), 7.80 (s,
1H, ArH), 7.93 (d, 1H, J = 2.1 Hz, ArH), 8.23–8.28 (m, 2H,
ArH), 8.40 (d, 1H, J = 8 Hz, ArH), 9.12 (s, 1H, ArH); MS:
m/e 357 (M + 1).
4-(9-methyl-9H-carbazol-3-yl)-6-(pyridin-2-yl)-pyrimidin-2-
amine (5o)
Yield: 65%; MP: 161–164°C; IR (KBr, cm−1): 3368, 3325,
1
3019, 1595, 1560, 1445, 1369, 1212, 1155, 1045, 932; H
NMR (200 MHz, DMSO-d6, δ in ppm): 3.87 (s, 3H, NCH3),
6.80 (s, 2H, NH2), 7.29 (t, 1H, J = 8 Hz, ArH), 7.48–7.58
(m, 4H, ArH), 7.61–7.73 (m, 2H, ArH), 7.80 (s, 1H, ArH),
8.25–8.38 (m, 2H, ArH), 8.40 (d, 1H, J = 8 Hz, ArH), 9.12
(s, 1H, ArH); MS: m/e 352 (M + 1).
Calculation of drug-likeness properties
e physicochemical properties such as molecular
weight, c Log P, HBA, HBD, polar surface area, molecular
volume and drug likeness of the synthesized compounds
Journal of Enzyme Inhibition and Medicinal Chemistry

Novel series of aminopyrimidine derivatives
7
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Acknowledgments
HVC grateful to Council of Scientific and Industrial
Research (CSIR), New Delhi, India for financial assistance
in the form of SRF.
Declaration of interest
e authors report no conflicts of interest.
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